Biomedical subjects

Martin Judex

Publications and source records attributed to Martin Judex.

6 recordsLinked to original sources

PubMed2006

CCL18/PARC stimulates hematopoiesis in long-term bone marrow cultures indirectly through its effect on monocytes.

Chemokines play a role in regulating hematopoietic stem cell function, including migration, proliferation, and retention. We investigated the involvement of CCL18 in the regulation of bone marrow hematopoiesis. Treatment of human long-term bone marrow cultures (LTBMCs) with CCL18 resulted in significant stimulation of hematopoiesis, as measured by the total number of hematopoietic cells and their committed progenitors produced in culture. Monocytes/macrophages, whose survival was almost doubled in the presence of CCL18 compared with controls, were the primary cells mediating this effect. Conditioned media from CCL18-treated mature monocytes fostered colony-promoting activity that increased the number of colonies formed by hematopoietic progenitor cells. Gene expression profiling of CCL18-stimulated monocytes demonstrated more than 200 differentially expressed genes, including those regulating apoptosis (caspase-8) and proliferation (IL-6, IL-15, stem cell factor [SCF]). Up-regulation of these cytokines was confirmed on the protein expression level. The contribution of SCF and IL-6 in CCL18-mediated stimulatory activity for hematopoiesis was confirmed by SCF- and IL-6-blocking antibodies that significantly inhibited the colony-promoting activity of CCL18-stimulated conditioned medium. In addition to the effect on monocytes, CCL18 facilitated the formation of the adherent layer in LTBMCs and increased the proliferation of stromal fibroblast-like cells.

Apoptosis↗

PubMed2006

Vertical arrays: microarrays of complex mixtures of nucleic acids.

Vertical arrays are microarrays that have complex mixtures of nucleic acids as array elements, and that are hybridized with single sequence probes. Like dot blots, many different experiments can be spotted on a single vertical array, allowing single genes to be compared across many conditions. Vertical arrays have two additional advantages over dot blots. First, they are printed on glass slides, allowing the use of low-volume, high-concentration hybridization reactions. Second, they can be made using low-complexity representations of the original nucleic acid population. This increases signal-to-noise relative to the usual use of dot blots, wherein the entire complexity of the population is usually spotted. Whereas standard microarrays achieve horizontal coverage of many genes and are repeated to cover many experiments, vertical arrays achieve vertical coverage of many experiments and are repeated to cover many genes. In cases where the number of genes is limited, but the number of experiments is very large, vertical arrays may be advantageous.

Animals↗

PubMed2005

Enhanced microarray performance using low complexity representations of the transcriptome.

Low abundance mRNAs are more difficult to examine using microarrays than high abundance mRNAs due to the effect of concentration on hybridization kinetics and signal-to-noise ratios. This report describes the use of low complexity representations (LCRs) of mRNA as the targets for cDNA microarrays. Individual sequences in LCRs are more highly represented than in the mRNA populations from which they are derived, leading to favorable hybridization kinetics. LCR targets permit the measurement of abundance changes that are difficult to measure using oligo(dT) priming for target synthesis. An oligo(dT)-primed target and three LCRs detect twice as many differentially regulated genes as could be detected by the oligo(dT)-primed target alone, in an experiment in which serum-starved fibroblasts responded to the reintroduction of serum. Thus, this target preparation strategy considerably increases the sensitivity of cDNA microarrays.

Cell Line↗

PubMed2004

Laser-mediated microdissection as a tool for molecular analysis in arthritis.

Most current approaches to the analysis of gene expression in arthritic tissue samples are based on RNA isolated either from cultured synovial cells or from synovial biopsies. However, this strategy does not distinguish between specific gene expression profiles of cells originating from separate tissue areas. Therefore, we established the combination of laser-mediated microdissection and RNA arbitrarily primed polymerase chain reaction (RAP-PCR) for differential display to analyze profiles of gene expression in histologically defined areas of arthritic tissue. Cryosections derived from synovial tissue were used to obtain cell samples from different tissue areas using a microbeam laser microscope. RNA was isolated and analyzed using nested RAP-PCR to generate a fingerprint of the expressed gene sequences. Differentially expressed bands were isolated, cloned, and sequenced. Differential expression of identified sequences was confirmed by in situ hybridization and immunohistochemistry.

Arthritis↗

PubMed2002

Local production of complement proteins in rheumatoid arthritis synovium.

OBJECTIVE: Complement has been repeatedly implicated in the pathogenesis of rheumatoid arthritis (RA) based on studies showing reduced levels of native complement components and increased levels of complement metabolites in plasma, synovial fluid (SF), and synovial tissue (ST) of RA patients. However, there is limited information on local production and activation of key factors of the complement cascade in RA synovium and their potential modulation by novel anticytokine therapies. This study was undertaken to characterize the expression of complement proteins and receptors in RA SF and ST. METHODS: Using in situ hybridization, immunohistochemistry, and Western blot techniques, we assessed the presence of complement proteins C3, factor B (FB), and C5b-9, as well as the expression of complement receptors C3aR and C5aR in rheumatoid synovium. C3 and FB levels in SF were determined by enzyme-linked immunosorbent assay. Functional assessment was performed by examining the effects of soluble tumor necrosis factor receptor (sTNFR) p55 gene transfer in the SCID mouse model of RA. RESULTS: Complement proteins and receptors could be localized in all RA synovial specimens, whereas in osteoarthritis (OA) synovium, only a few, single cells expressed complement proteins and receptors. No differences were noted in the concentration of C3 between RA and OA in SF; however, FB levels were markedly reduced in RA versus OA SF. In RA synovium, in contrast to OA synovium, local expression of complement factor and complement receptor messenger RNA was found throughout the various ST compartments, suggesting that activation of the complement cascade occurs in all parts of the rheumatoid synovium. Moreover, C5aR expression was up-regulated following overexpression of sTNFR p55 by adenovirus-based gene transfer. CONCLUSION: In summary, local complement production and activation may play an important role in RA, and specific modulation and inhibition of local complement production could be an attractive therapeutic target for RA.

Aged↗

PubMed2002

Transcription factor Egr-1 activates collagen expression in immortalized fibroblasts or fibrosarcoma cells.

Synovial fibroblasts from rheumatoid arthritis patients express elevated levels of the transcription factor Egr-1. The metabolic consequences of Egr-1 overexpression in fibroblasts are not known in detail. Therefore we searched for gene products that are differentially expressed in Egr-1(high) versus Egr-1(low) fibroblasts. Immortalized synovial fibroblasts were transfected with two different Egr-1 expression vectors. Expression of recombinant Egr-1 was confirmed by RT-PCR and immunoblots. Random arbitrarily primed PCR revealed that Egr-1 induces enhanced transcription levels of the alpha1 chain of type I collagen. Increased expression of the alpha2 (I) chain could also be observed. We found enhanced levels of type I collagen propeptide in supernatants and stronger signals of alpha2 (I) protein in extracts of the Egr-1(high) expressing clone versus controls. Additionally, Egr-1 was transiently expressed in fibrosarcoma cells. These cells showed a pronounced elevation of type I collagen (alpha1) transcripts as well. Moreover, we could demonstrate that Egr-1 induces transcription of other genes including type II collagen (alpha1) and plateled-derived growth factor beta1. These data suggest that upregulation of Egr-1 might contribute tofibrosis observed in rheumatoid arthritis synovium by activation of genes encoding the alpha1 and alpha2 chains of type I collagen.

Cloning, Molecular↗