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1.
Cancer Epidemiol ; 38(1): 85-92, 2014 Feb.
Article in English | MEDLINE | ID: mdl-24445140

ABSTRACT

The miR-451 was found to be frequently down-regulated in tumors, indicating that miR-451 could play an important role in carcinogenesis. This study uncovered the mechanism by which the miR-451 functions as a tumor suppressor. The target genes of miR-451 were determined using target gene prediction softwares. Then the miR-451 mimics were introduced into RKO and Hela cells respectively. The proliferation and invasion of cells were monitored by MTT, cell cycle and in vitro extracellular matrix invasion assays. Also the angiogenesis of HUVEC cells transfected with miR-451 mimics was examined. Subsequently, IL6R, a predicted target gene of miR-451, was studied by real time PCR, Western blotting, and siRNA technologies. The mRNA and protein levels of IL6R gene were found to be down-regulated in the RKO and Hela cells transfected with miR-451 mimics. Consequently, the cell proliferation was inhibited. Also, the invasion of RKO cells was suppressed. Furthermore, the angiogenesis of HUVEC cells transfected with miR-451 mimics was assayed and the decreased angiogenic ability was detected compared to the controls. All these results were validated by IL6R siRNA experiments. The IL6R gene is a target gene of miR-451. The miR-451 behaves as a tumor suppressor, probably by targeting the IL6R pathway.


Subject(s)
Colonic Neoplasms/genetics , MicroRNAs/genetics , Receptors, Interleukin-6/genetics , Uterine Cervical Neoplasms/genetics , Blotting, Western , Cell Line, Tumor , Cell Proliferation , Colonic Neoplasms/pathology , Down-Regulation , Female , Gene Expression Regulation, Neoplastic , HeLa Cells , Human Umbilical Vein Endothelial Cells , Humans , Neovascularization, Physiologic/genetics , Real-Time Polymerase Chain Reaction , Transfection , Uterine Cervical Neoplasms/pathology
2.
Anticancer Res ; 32(11): 4861-4, 2012 Nov.
Article in English | MEDLINE | ID: mdl-23155252

ABSTRACT

AIM: Our previous study showed that methylation of the mutL homolog 1 (MLH1) promoter may spread upstream from the Alu elements in intron 1. In this study, we investigated if the Alu methylation could also spread downstream. MATERIALS AND METHODS: Two colorectal cancer cell lines (RKO, SW48), and four colorectal and three gastric carcinomas [all Microsatellite Instability (MSI)-positive] were selected as cases. Normal colorectal and gastric mucosa and human peripheral blood, and one MSI-negative colorectal cancer case served as controls. After extraction of DNA, bisulfite genomic sequencing was used to analyze the methylation level of exon 2 and the adjacent part of intron 2 of the MLH1 gene. RESULTS: Exon 2 and the partial intron 2 exhibited a high level of methylation in controls. In contrast, demethylation in these regions was seen in gastrointestinal cancer. CONCLUSION: Exon 2 methylation is not likely to influence MLH1 gene expression. It seems that de-methylation of exon 2 and intron 2, in combination with intron 1, is associated with methylation spreading of the MLH1 promoter. The region around exon 2 could possibly bind proteins that regulate methylation, and therefore affect gene expression.


Subject(s)
Adaptor Proteins, Signal Transducing/genetics , DNA Methylation/genetics , Gastrointestinal Neoplasms/genetics , Gene Expression Regulation, Neoplastic/genetics , Nuclear Proteins/genetics , Adaptor Proteins, Signal Transducing/metabolism , Cell Line, Tumor , Exons , Gastrointestinal Neoplasms/metabolism , Gene Expression , Humans , MutL Protein Homolog 1 , Nuclear Proteins/metabolism , Polymerase Chain Reaction , Promoter Regions, Genetic/genetics
3.
Nat Cell Biol ; 14(2): 201-8, 2012 Jan 29.
Article in English | MEDLINE | ID: mdl-22286099

ABSTRACT

There are three prolyl hydroxylases (PHD1, 2 and 3) that regulate the hypoxia-inducible factors (HIFs), the master transcriptional regulators that respond to changes in intracellular O(2) tension. In high O(2) tension (normoxia) the PHDs hydroxylate two conserved proline residues on HIF-1α, which leads to binding of the von Hippel-Lindau (VHL) tumour suppressor, the recognition component of a ubiquitin-ligase complex, initiating HIF-1α ubiquitylation and degradation. However, it is not known whether PHDs and VHL act separately to exert their enzymatic activities on HIF-1α or as a multiprotein complex. Here we show that the tumour suppressor protein LIMD1 (LIM domain-containing protein) acts as a molecular scaffold, simultaneously binding the PHDs and VHL, thereby assembling a PHD-LIMD1-VHL protein complex and creating an enzymatic niche that enables efficient degradation of HIF-1α. Depletion of endogenous LIMD1 increases HIF-1α levels and transcriptional activity in both normoxia and hypoxia. Conversely, LIMD1 expression downregulates HIF-1 transcriptional activity in a manner depending on PHD and 26S proteasome activities. LIMD1 family member proteins Ajuba and WTIP also bind to VHL and PHDs 1 and 3, indicating that these LIM domain-containing proteins represent a previously unrecognized group of hypoxic regulators.


Subject(s)
Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Intracellular Signaling Peptides and Proteins/metabolism , LIM Domain Proteins/metabolism , Procollagen-Proline Dioxygenase/metabolism , Von Hippel-Lindau Tumor Suppressor Protein/metabolism , Cell Hypoxia , Cell Line, Tumor , HEK293 Cells , HeLa Cells , Humans , Hydroxylation , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Hypoxia-Inducible Factor-Proline Dioxygenases , Immunoblotting , Immunoprecipitation , Intracellular Signaling Peptides and Proteins/genetics , LIM Domain Proteins/genetics , Models, Biological , Polyubiquitin/metabolism , Procollagen-Proline Dioxygenase/genetics , Proteasome Endopeptidase Complex/metabolism , Protein Binding , RNA Interference , Transfection , Two-Hybrid System Techniques , Ubiquitination , Von Hippel-Lindau Tumor Suppressor Protein/genetics
4.
PLoS One ; 6(12): e28730, 2011.
Article in English | MEDLINE | ID: mdl-22194897

ABSTRACT

miRNAs have been found to repress gene expression at posttranscriptional level in cells. Studies have shown that expression of miRNAs is tissue-specific and developmental-stage-specific. The mechanism behind this could be explained by miRNA pathways. In this study, totally 54 miRNAs were analysed in 7 matched human foetal and adult organs (brain, colon, heart, kidney, liver, lung and spleen) using real-time PCR. Quantitative analysis showed that a big proportion of the 54 miRNAs have higher general expression in the organs of the foetal period than the adult period, with the exception of the heart. The miRNA gene promoter methylation level in the adult stages was higher than in the foetal stages. Moreover, there is a high general expression level of several miRNAs in both stages of brain, kidney, liver, lung and spleen, but not seen in colon and heart. Our results indicate that the miRNAs may play a bigger role in the foetal stage than the adult stage of brain, colon, kidney, liver, lung and spleen. The majority of the miRNAs analysed may play an important role in the growth and development of brain, kidney, liver, lung and spleen. However, a minority of the miRNAs may be functional in colon and heart.


Subject(s)
Fetus/metabolism , Gene Expression Regulation, Developmental , MicroRNAs/genetics , Organ Specificity/genetics , Adolescent , Adult , DNA Methylation/genetics , Female , Gene Expression Profiling , Humans , MicroRNAs/metabolism , Promoter Regions, Genetic/genetics
5.
PLoS One ; 6(10): e25913, 2011.
Article in English | MEDLINE | ID: mdl-22022465

ABSTRACT

The highly repetitive Alu retroelements are regarded as methylation centres in the genome. Methylation in the gene promoters could be spreading from them. Promoter methylation of MLH1 is frequently detected in cancers, but the underlying mechanism is unclear. The aim of this study is to understand whether the methylation in the Alu elements is associated with promoter methylation in the MLH1 gene. Bisulfite genomic sequencing was used to analyse the CpG sites of the 5' end (promoter, exon 1 and Alu-containing intron 1) of the MLH1 gene in colorectal cancer cells and tissues, and gastric cancer tissues. Hypomethylation in the Alu elements and hypermethylation in the promoters and the regions between the promoters and the Alu elements were detected in two cancer cell lines and seven cancer tissues. However, demethylation or hypomethylation of the MLH1 promoter and regions between promoter and the Alu elements, and hypermethylation in the Alu elements, were identified in the normal tissues. MLH1 promoter methylation may spread from Alu elements that are located in intron 1 of the MLH1 gene. The trans-acting elements binding to the mutation sites could play a role in the methylation spreading.


Subject(s)
Adaptor Proteins, Signal Transducing/genetics , Alu Elements/genetics , DNA Methylation/genetics , Gastrointestinal Neoplasms/genetics , Nuclear Proteins/genetics , Promoter Regions, Genetic , Base Sequence , Cell Line, Tumor , CpG Islands/genetics , DNA Mutational Analysis , Exons/genetics , Gastrointestinal Neoplasms/pathology , Genetic Testing , Humans , Immunohistochemistry , Introns/genetics , Molecular Sequence Data , MutL Protein Homolog 1
6.
Hepatol Res ; 40(8): 813-22, 2010 Aug.
Article in English | MEDLINE | ID: mdl-20649821

ABSTRACT

AIM: miRNAs have been found to regulate gene expression at a posttranscriptional level in cells. Studies have shown that expression of miRNAs is tissue-specific and developmental stage-specific. The mechanism behind this could be explained by miRNA pathways. METHODS: We introduce the identification of miRNAs from two human fetal liver cDNA libraries by a cloning protocol. The miRNAs detected were then analyzed in a chorionic villus tissue and four liver tissues using real-time polymerase chain reaction. RESULTS: After sequencing and database searching, a total of 42 miRNAs in two fetal livers were detected. Quantitative analysis showed that they have higher general expression in the livers of the fetal period than the adult period, and furthermore the expression levels of the miRNAs during the fetal period were dynamically changed. CONCLUSION: Our results indicate that a special group of miRNAs may play an important role in human fetal liver development, while their roles in the adult livers are limited.

7.
Cancer Epidemiol ; 34(1): 93-5, 2010 Feb.
Article in English | MEDLINE | ID: mdl-20060799

ABSTRACT

The -93 SNP of MLH1 gene is associated with MLH1 gene methylation in endometrial and colorectal cancers. We undertook luciferase reporter assay and electrophoretic mobility shift assay (EMSA) to test whether the -93 SNP affects the MLH1 gene expression. The luciferase activity for -93A plasmid is significantly lower than -93G plasmid. In EMSA experiments, the -93A and -93G probes have different binding affinity to nuclear proteins of JEG3 cells. Our data indicate that -93 SNP affects MLH1 gene expression by altering protein binding to the promoter of MLH1 gene.


Subject(s)
Adaptor Proteins, Signal Transducing/genetics , Gene Expression Regulation , Nuclear Proteins/genetics , Polymorphism, Single Nucleotide , Promoter Regions, Genetic , Adaptor Proteins, Signal Transducing/metabolism , Colorectal Neoplasms/genetics , DNA Methylation , DNA-Binding Proteins/metabolism , Endometrial Neoplasms/genetics , Female , Humans , Luciferases/analysis , Luciferases/genetics , Male , MutL Protein Homolog 1 , Nuclear Proteins/metabolism
8.
PLoS One ; 4(10): e7594, 2009 Oct 26.
Article in English | MEDLINE | ID: mdl-19855840

ABSTRACT

BACKGROUND: miRNAs are 17-25 nucleotides long RNA molecules that have been found to regulate gene expression in human cells. There are studies showing that different groups of miRNAs are involved in development of different tissues. In hepatocytes there are reported particular types of miRNAs that regulate gene expression. METHODS: We established a human fetal liver cDNA library by a modified cloning protocol. Then plasmid isolation from the colonies was performed. After sequencing and database searching, the miRNAs were recognized. RT-PCR and sequencing were carried out to validate the miRNAs detected. Real-time PCR was used to analyze the expression of each miRNA. RESULTS: One novel miRNA was discovered, together with another 35 previously-known miRNAs in the fetal liver. Some of them existed in variants. The miRNAs identified were validated by RT-PCR and sequencing. Quantitative analysis showed that they have variable expression. CONCLUSION: Our results indicate that a special group of miRNAs may play an important role in fetal liver development in a synergistic manner.


Subject(s)
Genetic Techniques , Liver/embryology , Liver/metabolism , MicroRNAs/analysis , Reverse Transcriptase Polymerase Chain Reaction/methods , Base Sequence , Cells, Cultured , Female , Fetus/metabolism , Gene Library , Humans , Liver/chemistry , MicroRNAs/genetics , Molecular Sequence Data , Nucleic Acid Conformation , Sequence Homology, Nucleic Acid
9.
Acta Vet Scand ; 50: 44, 2008 Nov 04.
Article in English | MEDLINE | ID: mdl-18983656

ABSTRACT

One of the prerequisite for developing DNA vaccines for horses are vectors that are efficiently expressed in horse cells. We have analysed the ectopic expression of the human serum albumin gene in primary horse cells from different tissues. The vectors used are of pcDNA and pUC origin and include the cytomegalovirus (CMV) promoter. The pUC vectors contain CMV intron A whereas the pcDNA vectors do not. Insertion of intron A diminished the expression from the pcDNA vectors whereas insertion of a Kozak sequence upstream of the gene in two types of pUC vectors increased significantly the in vitro expression in primary horse cells derived from skin, lung, duodenum and kidney. We report for the first time the significance of full consensus Kozak sequences for protein expression in horse cells in vitro.


Subject(s)
Horses/immunology , Vaccines, DNA/genetics , Animals , Base Sequence , Blotting, Western/veterinary , Cell Line , Cytomegalovirus/genetics , Genetic Vectors/genetics , Genetic Vectors/immunology , Interferon alpha-2 , Interferon-alpha/biosynthesis , Interferon-alpha/genetics , Interferon-alpha/immunology , Plasmids/genetics , Recombinant Proteins , Serum Albumin/biosynthesis , Serum Albumin/genetics , Transfection/methods , Transfection/veterinary , Vaccines, DNA/immunology
10.
J Virol ; 81(8): 4052-7, 2007 Apr.
Article in English | MEDLINE | ID: mdl-17287273

ABSTRACT

Maedi-visna virus (MVV) is a lentivirus of sheep causing chronic inflammatory disease of the lungs (maedi) and the nervous system (visna). We have previously shown that a duplicated sequence in the long terminal repeat (LTR) of MVV is a determinant of cell tropism. Here, we demonstrate that deletion of a CAAAT sequence from either one of the repeats resulted in poor virus growth in sheep choroid plexus cells. A duplication in the LTR encompassing the CAAAT sequence was found in four neurological field cases that were sequenced, but no duplication was present in the LTRs from seven maedi cases; one maedi isolate was mixed. These results indicate that the duplication in the LTR is associated with neurovirulence.


Subject(s)
Choroid Plexus/virology , Terminal Repeat Sequences/genetics , Visna-maedi virus/physiology , Visna-maedi virus/pathogenicity , Animals , Base Sequence , Cells, Cultured , Choroid Plexus/cytology , Molecular Sequence Data , Pneumonia, Progressive Interstitial, of Sheep/virology , Sequence Deletion , Sheep, Domestic , Visna/virology , Visna-maedi virus/genetics , Visna-maedi virus/isolation & purification
11.
Cancer Genomics Proteomics ; 3(3-4): 137-146, 2006.
Article in English | MEDLINE | ID: mdl-31394692

ABSTRACT

The genome of breast tumour cells is considered to be unstable, as reflected by multiple chromosomal and gene abnormalities. The molecular mechanism of genomic instability progression in breast cancer is poorly understood, but recent data suggest that mutated or overexpressed proteins affect the genome in several ways, including an abnormal number of centrosomes, inefficient DNA repair and unwanted telomere maintenance. Among these proteins are p53, Brca1, Brca2, Aurora kinase A, Myc and telomerase. The involved molecular networks include co-regulation with cell cycle checkpoints. p53 has been relatively well studied and is considered to be a guardian of the genome integrity. Myc seems to affect tumour pathogenesis in several ways, including increased proliferation and immortalisation of the cancer cells and induction of genomic instability. Aurora kinase A has been shown to control the centrosome number of cells and the segregation of the correct chromosomes to the daughter cells during mitosis. Genomic instability is high in some hereditary breast cancer, particularly in tumours of Brca1- and Brca2-mutation carriers, a finding that is in line with their role in DNA repair.

12.
Oncogene ; 24(28): 4540-8, 2005 Jun 30.
Article in English | MEDLINE | ID: mdl-15856022

ABSTRACT

An emerging paradigm holds that loss of negative signalling to receptor tyrosine kinases (RTKs) is permissive for their oncogenic activity. Herein, we have addressed tumor suppression by RALT/MIG-6, a transcriptionally controlled feedback inhibitor of ErbB RTKs, in breast cancer cells. Knockdown of RALT expression by RNAi enhanced the EGF-dependent proliferation of normal breast epithelial cells, indicating that loss of RALT signalling in breast epithelium may represent an advantageous condition during ErbB-driven tumorigenesis. Although mutational inactivation of the RALT gene was not detected in human breast carcinomas, RALT mRNA and protein expression was strongly and selectively reduced in ERBB2-amplified breast cancer cell lines. Reconstitution of RALT expression in ERBB2-amplified SKBr-3 and BT474 cells inhibited ErbB-2-dependent mitogenic signalling and counteracted the ability of ErbB ligands to promote resistance to the ErbB-2-targeting drug Herceptin. Thus, loss of RALT expression cooperates with ERBB2 gene amplification to drive full oncogenic signalling by the ErbB-2 receptor. Moreover, loss of RALT signalling may adversely affect tumor responses to ErbB-2-targeting agents.


Subject(s)
Adaptor Proteins, Signal Transducing/genetics , Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/pharmacology , Breast Neoplasms/genetics , Carcinoma/genetics , Receptor, ErbB-2/genetics , Adaptor Proteins, Signal Transducing/drug effects , Adaptor Proteins, Signal Transducing/metabolism , Antibodies, Monoclonal, Humanized , Breast Neoplasms/drug therapy , Breast Neoplasms/pathology , Carcinoma/pathology , Cell Line, Tumor , DNA Mutational Analysis , Drug Resistance, Neoplasm/genetics , Epidermal Growth Factor/pharmacology , Extracellular Signal-Regulated MAP Kinases/metabolism , Female , Gene Expression Regulation, Neoplastic , Humans , Loss of Heterozygosity , Receptor, ErbB-2/metabolism , Signal Transduction , Trastuzumab , Tumor Suppressor Proteins
13.
Cancer Genomics Proteomics ; 2(4): 247-253, 2005.
Article in English | MEDLINE | ID: mdl-31394713

ABSTRACT

The short arm of chromosome 3 is frequently altered in human cancers of different tissue origin. Certain regions on the chromosome arm 3p have been defined by deletion studies in human cancer cells and tissues. Also, regions at 3p are eliminated in microcell hybrids parallel to increased tumorigenicity in immunosuppressed mice. We have analysed chromosome instability and several genes involved in tumor pathogenesis at 3p, such as VHL, RIITGFB, CATNB, MLH1 and FHIT. By studying eleven tumor types, we have shown that the importance of CER1 (common eliminated region 1) transgresses tissue specificity. Comparative studies on losses of VHL, FHIT/FRA3B and CER1 show that the CER1 region is preferably lost in human tumors. Alterations of FHIT are associated with reduced survival of breast and colon cancer patients. The FHIT gene is located at the constitutive fragile region, FRA3B. Chromosome 3, particularly FHIT, is unstable in breast cancer patients who have germ line mutation in the BRCA2 gene. The chromosome instability in BRCA2 tumors reflects the DNA repair mechanism of the gene product Brca2. It can be concluded that our results reflect a synergism of tumor suppressor gene (TSG) losses at the chromosome 3p region in relation to the biological behavior of tumor cells and tumor pathogenesis.

14.
Genes Chromosomes Cancer ; 41(3): 232-42, 2004 Nov.
Article in English | MEDLINE | ID: mdl-15334546

ABSTRACT

A human chromosomal segment regularly lost during tumor formation of microcell hybrids in SCID mice has been mapped to 3p21.3. This segment, called chromosome 3 common eliminated region 1 (C3CER1, also referred to as CER1), may harbor multiple tumor-suppressor genes. Because it was found that similar regions were eliminated in an inter- and intraspecies system and in two tumor types (mouse fibrosarcoma and human renal cell carcinoma), we hypothesized that the importance of C3CER1 would transgress tissue specificity, that is, it could occur in tumors derived from multiple tissues. To evaluate the loss of C3CER1 in various human tumor types, we conducted loss of heterozygosity (LOH) analysis of 576 human solid tumors from 10 different tissues and compared the frequency of deletion in the C3CER1 area to that in two other regions on 3p: the FHIT/FRA3B region, at 3p14.2, and the VHL region, at 3p25.3. Deletions were detected in the C3CER1 region in 83% of informative tumors. Half (47%) the LOH-positive tumors showed LOH at all informative markers, indicating a large deletion. The other half (53%) had a discontinuous LOH pattern, suggesting interstitial deletions or breakpoints. The proportion of tumors with C3CER1 deletions was high in all tumor types investigated, ranging from 70% to 94%, except for the soft-tissue sarcomas (40%). In the VHL and FHIT regions, deletions were observed in 73% and 43%, respectively, of the tumors. Of the three 3p regions analyzed, the highest deletion frequency was observed in the C3CER1 region. Furthermore, we demonstrated that the interstitial deletions including C3CER1 prevail over 3p14.2-pter losses in solid tumors.


Subject(s)
Chromosomes, Human, Pair 3/ultrastructure , Gene Deletion , Neoplasms/genetics , Acid Anhydride Hydrolases/genetics , Biomarkers, Tumor , Cell Line, Tumor , Female , Humans , Loss of Heterozygosity , Male , Models, Genetic , Neoplasm Proteins/genetics , Neoplasms/metabolism , Polymerase Chain Reaction , Tumor Suppressor Proteins/genetics , Ubiquitin-Protein Ligases/genetics , Von Hippel-Lindau Tumor Suppressor Protein
15.
Drugs Today (Barc) ; 40(12): 991-1002, 2004 Dec.
Article in English | MEDLINE | ID: mdl-15645010

ABSTRACT

Breast cancer has all the hallmarks of a multistep genetic disease. Somatic and germ-line mutations have been described in several tumor suppressor genes, and oncogenes are found to be amplified. Genes in the ATM-CHK2-TP53 cell-cycle checkpoint pathway are mutated in relation to breast cancer, particularly TP53 at the somatic level. Germ-line mutations in BRCA1 and BRCA2, in which DNA repair function is interrupted, account for the majority of familial breast cancers. The mechanism behind the frequent instability of the genomes of breast cancer cells has been poorly understood, but recent functional findings on oncogenes and tumor suppressor genes have provided substantial information on the matter. Some recent developments in drug therapy are based on molecular and genomic findings about breast cancer pathogenesis.


Subject(s)
Breast Neoplasms/genetics , Genomic Instability/genetics , Animals , Breast Neoplasms/physiopathology , Female , Genes, Tumor Suppressor/physiology , Humans
16.
J Immunol ; 171(4): 1657-66, 2003 Aug 15.
Article in English | MEDLINE | ID: mdl-12902463

ABSTRACT

In the present study we demonstrate that all CD4(+) T cells in human tonsil expressing the Th2-selective receptor chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) also 1) express high levels of CXCR5, and 2) display a transitional CD45RA/RO phenotype and consistently do not produce significant amounts of cytokines when immediately analyzed ex vivo. Hence, they represent precursors of Th2 effector cells, a conclusion confirmed by their robust production of IL-4, IL-5, and IL-13, but not IFN-gamma, after in vitro activation. CD4(+) T cells, which express only intermediate levels of CXCR5, instead develop into IFN-gamma-producing cells under identical culture conditions, thus establishing a correlation between relative levels of CXCR5 expression and the acquired cytokine profile. Because CXCR5 is critically involved in follicular localization, the results suggest that these CRTH2(+) Th2 cells preferentially develop their cytokine-producing phenotype within germinal centers (GCs), whereas extrafollicular differentiation instead promotes Th1 development. In support for this proposal, we show that T cells with an intermediate expression of CXCR5 can be forced to also produce IL-4 and IL-13 if cultured with allogenic GC B cells. Finally, we demonstrate that the previously described CD57(+) GC T cells also express high levels of CXCR5 but instead of comprising a Th2 precursor, they represent anergized T cells. Taken together, these data suggest that GCs and B cells regulate CD4(+) T cell differentiation in a finely tuned fashion, either by promoting differentiation of Th2 cells, which apparently leave the lymphoid tissue before evolving a cytokine-producing phenotype, or by furnishing T cell unresponsiveness.


Subject(s)
Cell Differentiation/immunology , Germinal Center/cytology , Germinal Center/immunology , Receptors, Prostaglandin , Th2 Cells/cytology , B-Lymphocyte Subsets/immunology , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD57 Antigens/biosynthesis , Cells, Cultured , Chemokines, CXC/metabolism , Child , Clonal Anergy , Coculture Techniques , Cytokines/biosynthesis , Germinal Center/metabolism , Humans , Immunologic Memory , Immunophenotyping , Interferon-gamma/biosynthesis , Interleukin-13/biosynthesis , Interleukin-4/biosynthesis , Lymphocyte Activation/immunology , Palatine Tonsil , Receptors, Antigen, T-Cell/physiology , Receptors, CXCR5 , Receptors, Chemokine , Receptors, Cytokine/biosynthesis , Receptors, Immunologic/biosynthesis , Stem Cells/cytology , Stem Cells/immunology , Stem Cells/metabolism , T-Lymphocytes, Helper-Inducer/cytology , T-Lymphocytes, Helper-Inducer/immunology , T-Lymphocytes, Helper-Inducer/metabolism , Th2 Cells/immunology , Th2 Cells/metabolism
17.
Blood ; 101(6): 2307-13, 2003 Mar 15.
Article in English | MEDLINE | ID: mdl-12411303

ABSTRACT

A number of studies have implicated a role for the cell surface glycoprotein CD44 in several biologic events, such as lymphopoiesis, homing, lymphocyte activation, and apoptosis. We have earlier reported that signaling via CD44 on naive B cells in addition to B-cell receptor (BCR) and CD40 engagement generated a germinal center-like phenotype. To further characterize the global role of CD44 in B differentiation, we examined the expression profile of human B cells cultured in vitro in the presence or absence of CD44 ligation, together with anti-immunoglobulin (anti-Ig) and anti-CD40 antibodies. The data sets derived from DNA microarrays were analyzed using a novel statistical analysis scheme created to retrieve the most likely expression pattern of CD44 ligation. Our results show that genes such as interleukin-6 (IL-6), IL-1alpha, and beta(2)-adrenergic receptor (beta(2)-AR) were specifically up-regulated by CD44 ligation, suggesting a novel role for CD44 in immunoregulation and inflammation.


Subject(s)
B-Lymphocytes/chemistry , B-Lymphocytes/immunology , Hyaluronan Receptors/immunology , Immunity , Inflammation , Oligonucleotide Array Sequence Analysis , RNA, Messenger/analysis , Antibodies/pharmacology , Antibodies, Anti-Idiotypic/pharmacology , CD40 Antigens/immunology , Cells, Cultured , Gene Expression Profiling , Gene Expression Regulation/drug effects , Humans , Hyaluronan Receptors/metabolism , Interleukin-1/analysis , Interleukin-1/genetics , Interleukin-6/analysis , Interleukin-6/genetics , Receptors, Adrenergic, beta-2/genetics
18.
Breast Cancer Res ; 4(3): R4, 2002.
Article in English | MEDLINE | ID: mdl-12052256

ABSTRACT

BACKGROUND: Mutations in the CHK2 gene at chromosome 22q12.1 have been reported in families with Li-Fraumeni syndrome. Chk2 is an effector kinase that is activated in response to DNA damage and is involved in cell-cycle pathways and p53 pathways. METHODS: We screened 139 breast tumors for loss of heterozygosity at chromosome 22q, using seven microsatellite markers, and screened 119 breast tumors with single-strand conformation polymorphism and DNA sequencing for mutations in the CHK2 gene. RESULTS: Seventy-four of 139 sporadic breast tumors (53%) show loss of heterozygosity with at least one marker. These samples and 45 tumors from individuals carrying the BRCA2 999del5 mutation were screened for mutations in the CHK2 gene. In addition to putative polymorphic regions in short mononucleotide repeats in a non-coding exon and intron 2, a germ line variant (T59K) in the first coding exon was detected. On screening 1172 cancer patients for the T59K sequence variant, it was detected in a total of four breast-cancer patients, two colon-cancer patients, one stomach-cancer patient and one ovary-cancer patient, but not in 452 healthy individuals. A tumor-specific 5' splice site mutation at site +3 in intron 8 (TTgt [a --> c]atg) was also detected. CONCLUSION: We conclude that somatic CHK2 mutations are rare in breast cancer, but our results suggest a tumor suppressor function for CHK2 in a small proportion of breast tumors. Furthermore, our results suggest that the T59K CHK2 sequence variant is a low-penetrance allele with respect to tumor growth.


Subject(s)
Breast Neoplasms/genetics , Chromosomes, Human, Pair 22 , Loss of Heterozygosity , Mutation/genetics , Protein Kinases/genetics , Protein Serine-Threonine Kinases , Adult , Checkpoint Kinase 2 , Colonic Neoplasms/genetics , Female , Genes, BRCA2 , Heterozygote , Humans , Male , Middle Aged , Ovarian Neoplasms/genetics , Point Mutation/genetics , Polymorphism, Genetic , Stomach Neoplasms/genetics , Thyroid Neoplasms/genetics
19.
Anticancer Res ; 22(6A): 3205-12, 2002.
Article in English | MEDLINE | ID: mdl-12530066

ABSTRACT

BACKGROUND: Genomic alterations and abnormal expression of the FHIT gene have been reported for a number of cancers. FHIT encompasses FRA3B, the most common fragile site in the human genome, and is suggested to be a candidate tumour suppressor gene. MATERIALS AND METHODS: We analysed and compared the loss of heterozygosity (LOH) pattern in 397 solid human tumours from 9 different locations, using four polymorphic microsatellite markers within the gene (D3S1234, D3S1300, D3S2757 and D3S4260), and two markers (D3S1313 and D3S1600) flanking the gene. In addition, we tested whether there was an association between FHIT LOH and overall patient survival in colorectal cancer. RESULTS: LOH at the FHIT gene affecting at least one of the investigated markers was detected in 166 out of 332 informative tumours, or 50%. The highest detected LOH was in lung tumours (66%) while the lowest was in thyroid and endometrium tumours, (30% and 31%, respectively). Breakpoints were found inside the gene in all tumour types in 12-80% of the tumours with FHIT LOH depending on tumour type, and up to 41% could additionally be located adjacent to the 3' or 5' end of the FHIT gene. Thus we were able to locate breakpoints within or in the vicinity of the FHIT gene in 25-100% of different tumours with LOH. Although not statistically significant, we observed a trend towards a poorer survival of patients with FHIT LOH versus those with retention of heterozygosity. CONCLUSION: Based on our results, LOH of the FHIT gene is a common event in all tumour types analysed with a possible association with poorer survival in colorectal cancer patients. LOH at all markers analysed was, in most of the tumour types, a more common pattern of alterations than breakpoints.


Subject(s)
Acid Anhydride Hydrolases , Colorectal Neoplasms/genetics , Loss of Heterozygosity , Neoplasm Proteins/genetics , Neoplasms/genetics , Colorectal Neoplasms/mortality , Female , Genetic Markers/genetics , Humans , Male , Survival Rate
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