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Biomedical subjects

Ulf Müller-Ladner

Publications and source records attributed to Ulf Müller-Ladner.

At least 37 records · Page 2Linked to original sources
PubMed2005

The -2518 promotor polymorphism in the MCP-1 gene is associated with systemic sclerosis.

Factors influencing the initiation or progression of sclerosis in patients with systemic sclerosis (SSc) are poorly understood. Monocyte chemotactic protein-1 (MCP-1) is a potent chemokine, which is upregulated in fibroblasts during development of sclerosis. In this study, we investigated the frequency of the functional -2518G MCP-1 promoter polymorphism in 18 patients with SSc and 139 healthy controls. In the lesional skin of the same SSc patients, expression of MCP-1 protein was examined by immunohistochemistry. To investigate a genotype/phenotype correlation, basal as well as tumor necrosis factor (TNF)-induced MCP-1 expression was analyzed in fibroblasts isolated from the skin of SSc patients with different MCP-1 genotypes by quantitative RT-PCR and ELISA. Genotyping for the -2518 (A/G) MCP-1 promotor polymorphism showed that GG homozygotes were significantly more frequent in patients with SSc than in controls (28%vs 6%). Results of immunohistochemistry revealed that MCP-1 was expressed in keratinocytes, infiltrating inflammatory cells, fibroblasts, and endothelial cells in scleroderma skin, whereas normal control skin showed no MCP-1 expression. MCP-1 expression in fibroblasts from GG-homozygote individuals tended to be stronger as compared to AG or AA genotypes. Furthermore, basal as well as TNF-induced MCP-1 expression of fibroblasts isolated from a GG-homozygote SSc patient was significantly higher than MCP-1 expression of fibroblasts isolated from heterozygote or AA-homozygote donors. The A -2518G polymorphism of the MCP-1 gene appears to affect MCP-1 expression of skin fibroblasts of patients with SSc. In accordance, the G/G genotype may predispose patients to SSc.

Adult↗
PubMed2004

[Bronchobiliary fistula associated with combined abscess of lung and liver].

HISTORY AND PHYSICAL EXAMINATION: A 43-year-old, cachectic, alcohol-addicted patient was admitted to the hospital due to hemoptysis and fever of up to 40 degrees C for the last 4 days. Physical examination revealed pleural rubs in the right lower lung and a diffuse pain in the right upper quadrant of the abdomen. The right upper quadrant of the abdomen also showed scars from several surgical interventions on the small intestine, the liver and the biliary tract and deformed caudal ribs due to an accident 23 years earlier. DIAGNOSTICS, THERAPY, AND CLINICAL COURSE: Chest X-ray, ultrasound and computed tomography (CT) showed abscess cavities in lung and liver communicating through the diaphragm. Under antibiotic therapy the abscess was drained. Cultures showed Klebsiella pneumoniae and Streptococcus viridans. Continuous lavage of the abscess cavities and antibiotic therapy led to an improvement in parameters of inflammation and clinical status. Imaging after 10 days of treatment showed a contrast enrichment in the abscess cavities and a bronchobiliary fistula accounting for temporary biliptysis. The drainage was removed after 29 days, when similar imaging revealed no further sign of an abscess cavity and a previous CT follow-up had proven a distinct decrease of the abscesses. CONCLUSION: Bronchobiliary fistulas are very rare. In most cases they are caused by hepatic or subphrenic abscesses, resulting from different conditions. The development of an bronchobiliary fistula originating from a Klebsiella pneumoniae pneumonia, as indicated by this report, has not been described so far. In the patient presented here, treatment was achieved due to continuous drainage despite the large extent of the abscess.

Adult↗
PubMed2004

Uncontrolled expression of vascular endothelial growth factor and its receptors leads to insufficient skin angiogenesis in patients with systemic sclerosis.

Systemic sclerosis (SSc) skin lesions are characterized by disturbed vessel morphology with enlarged capillaries and an overall reduction in capillary density, suggesting a deregulated, insufficient angiogenic response. It has been postulated that this phenomenon is due to reduced expression of the potent angiogenic factor vascular endothelial growth factor (VEGF). In contrast to this hypothesis, we demonstrate that the expression of both VEGF and its receptors VEGFR-1 and VEGFR-2 is dramatically upregulated in skin specimens of SSc patients throughout different disease stages. Interestingly, upregulation of VEGF was not mediated by hypoxia-inducible transcription factor-1 (HIF-1) as indicated by only a weak expression of the oxygen-sensitive alpha-subunit of HIF-1 in the skin of SSc patients. This was unexpected on measuring low Po2 values in the SSc skin by using a polarographic oxygen microelectrode system. Considering our observation that PDGF and IL-1beta costimulated VEGF expression, we propose that chronic and uncontrolled VEGF upregulation that is mediated by an orchestrated expression of cytokines rather than VEGF downregulation is the cause of the disturbed vessel morphology in the skin of SSc patients. Consequently, for therapeutic approaches aiming to improve tissue perfusion in these patients, a controlled expression and timely termination of VEGF signaling appears to be crucial for success of proangiogenic therapies.

Cell Hypoxia↗
PubMed2004

Ribozymes that inhibit the production of matrix metalloproteinase 1 reduce the invasiveness of rheumatoid arthritis synovial fibroblasts.

OBJECTIVE: To investigate whether retroviral gene transfer of ribozymes targeting matrix metalloproteinase 1 (MMP-1) inhibits the production of MMP-1 in rheumatoid arthritis synovial fibroblasts (RASFs) and reduces the invasiveness of these cells in vivo. METHODS: MMP-1-specific ribozymes (RzMMP-1) were designed and cloned into the pLNSX retroviral vector. Cleavage of MMP-1 was determined in vitro, and the most effective ribozyme was selected for further investigation. RASFs were transduced with replication-deficient viruses carrying RzMMP-1 or with empty viruses (mock). Quantitative polymerase chain reaction with cleavage site-spanning fluorescent probes was used to measure the levels of MMP-1, MMP-9, and MMP-13 messenger RNA. In addition, protein levels of MMP-1 in cell culture supernatants were determined by enzyme linked immunosorbent assay. The effects of stimulation with lipopolysaccharide (LPS) and tumor necrosis factor alpha (TNFalpha) on the production of MMP-1 were assessed accordingly. The invasiveness of RzMMP-1-transduced, mock-transduced, and untransduced RASFs was analyzed in the SCID mouse in vivo model of RA. RESULTS: Transduction of RASFs with RzMMP-1 significantly decreased the production of MMP-1 in RASFs without affecting other MMPs, such as MMP-9 and MMP-13. RzMMP-1 not only reduced the spontaneous production of MMP-1, but also prevented the LPS- and TNFalpha-induced increase in MMP-1 production. Inhibition of MMP-1 was maintained for at least 2 months and was accompanied by a significant reduction of the invasiveness of RASFs in the SCID mouse model of RA. CONCLUSION: Intracellular expression of ribozymes constitutes a feasible tool for inhibiting the production of matrix-degrading enzymes. Inhibition of MMP-1 alone results in a significant reduction of cartilage invasion by RASFs.

Animals↗
PubMed2004

In vivo imaging of protease activity in arthritis: a novel approach for monitoring treatment response.

OBJECTIVE: Sensitive noninvasive strategies for monitoring treatment response in rheumatoid arthritis (RA) would be valuable for facilitating appropriate therapy and dosing, evaluating clinical outcome, and developing more effective drugs. Because different proteases are highly up-regulated in RA and contribute significantly to joint destruction, in the present study we investigated whether such enzymes are suitable in vivo imaging biomarkers for early evaluation of treatment response in a murine model of RA. METHODS: Using a protease-activated near-infrared fluorescence (NIRF) imaging "smart" probe, we examined the presence and distribution of fluorescence in arthritic joints of mice with collagen-induced arthritis by both noninvasive fluorescence imaging and histology. Proteases that target the Lys-Lys cleavage site, including cathepsin B, activate probe fluorescence. Treatment monitoring data were obtained following methotrexate (MTX) therapy. RESULTS: Twenty-four hours after intravenous injection of the protease sensor, affected toes and paws of arthritic mice showed significantly higher fluorescence intensity than did toes and paws of healthy mice. Fluorescence from the protease probe and cathepsin B antibody histologic staining were localized in the vast majority of cells in the inflamed synovium. In arthritic animals treated with MTX (35 mg of MTX/kg 48 hours prior to probe injection), a significantly lower fluorescent signal (inflamed paws 50%, inflamed toes 70%) was observed as compared with untreated arthritic animals. CONCLUSION: Protease-activated NIRF probes are sensitive means of imaging the presence of target enzymes in arthritic joints and can be used for early monitoring of treatment response to antirheumatic drugs such as MTX.

Animals↗
PubMed2004

Cooperation of Ras- and c-Myc-dependent pathways in regulating the growth and invasiveness of synovial fibroblasts in rheumatoid arthritis.

OBJECTIVE: To study the specific contribution of MAP kinase activator c-Raf-1 and one of its downstream transcription factors, c-Myc, to the growth and invasive behavior of rheumatoid arthritis synovial fibroblasts (RASFs). METHODS: RASFs were transduced with retroviral constructs expressing dominant-negative mutants of c-Raf-1 or c-Myc (DN c-Raf-1 or DN c-Myc, respectively) or with the mock vector. The expression of wild-type and mutant proteins was confirmed by Western blotting. Growth curves of RASFs were recorded, and apoptosis was measured by flow cytometry. Invasiveness of RASFs was assessed in the SCID mouse model of RA. Immunohistochemistry was used to study the effects of DN c-Raf-1 on phosphorylated c-Jun and matrix metalloproteinase 1 (MMP-1) in RASFs implanted into SCID mice. The phosphorylation of ERK and JNK in DN c-Raf-1- and mock-transduced RASFs was determined in vitro by Western blotting. The levels of MMPs in these cells were measured by quantitative polymerase chain reaction (PCR). RESULTS: Neither DN c-Raf-1 alone nor DN c-Myc alone significantly altered proliferation or apoptosis of RASFs, but both mutants together rapidly induced apoptosis. Inhibition of c-Raf-1 or c-Myc significantly reduced the invasiveness of RASFs in the SCID mouse model. DN c-Raf-1 decreased the phosphorylation of ERK and JNK in vitro and reduced the in vivo expression of phosphorylated c-Jun as well as the expression of disease-relevant MMPs. As determined by quantitative PCR, the inhibition was most pronounced for MMP-1 and MMP-3. CONCLUSION: The data demonstrate that Ras- and c-Myc-dependent signaling events cooperate to regulate the growth and invasiveness of RASFs. Targeting of both c-Raf-1 and c-Myc may constitute an interesting therapeutic approach in RA.

Animals↗
PubMed2004

Targeted gene therapy: frontiers in the development of 'smart drugs'.

Chronic diseases, particularly malignancies and immune-mediated inflammatory diseases (IMIDs), are a challenging frontier for clinical diagnosis and treatment, as well as for biomedical research. Current treatment regimens are frequently insufficient and thus new treatment strategies are needed. Novel therapies for disabling such diseases should provide improvements with respect to safety, efficacy and cost. To fulfill these three key criteria, recent research efforts have focused on the development of 'smart drugs'. This review highlights some examples of the rapidly expanding possibilities that current biotechnology has to offer in the development of novel therapeutic strategies for complex diseases such as IMIDs. Special attention is given to advances in, and limitations of, controlled and targeted gene product application in inflammatory diseases.

Antibodies, Monoclonal↗
PubMed2004

Functional genomics of fibroblasts.

PURPOSE OF REVIEW: Successful analysis of the pathophysiology of rheumatoid arthritis requires the functional understanding of interactions between different cell types and the cell matrix, intracellular signaling pathways, as well as between cartilage, bone, and synovium in rheumatoid arthritis. During the review period, molecular biology has provided and used a growing number of tools to screen the genome such as gene and protein chips, haplotype analysis, and single nucleotide polymorphism analysis, resulting in various novel findings with considerable impact on the overall understanding of rheumatoid arthritis. RECENT FINDINGS: Key issues that have been addressed and elucidated by numerous research groups are the regulation and modulation of synovial fibroblast metabolism and activation by proinflammatory cytokines and chemokines. In addition, examination of adhesion processes and neoangiogenesis has revealed new insights into the interaction network between rheumatoid synovial fibroblasts and the surrounding matrix and cells. Finally, a more detailed view of activation of these fibroblasts has been obtained by analysis of the molecular balance between cellular activation and regulation of apoptosis. SUMMARY: Although high throughput molecular analysis methods provided an ample amount of novel data, it needs to be stressed that a one-method approach of gene expression (eg, by array analysis) is not sufficient to validate the gene/gene product as a new therapeutic target. Therefore, the next steps are the so-called functional genomics or functionomics, which intend to reveal relations between the obtained data and to unveil their interactions for a better understanding of the pathogenesis and the mechanisms that are operative in rheumatoid arthritis.

Apoptosis↗
PubMed2004

Bucillamine induces the synthesis of vascular endothelial growth factor dose-dependently in systemic sclerosis fibroblasts via nuclear factor-kappaB and simian virus 40 promoter factor 1 pathways.

The pathogenesis of systemic sclerosis (SSc) is characterized by activation of the immune system, impaired angiogenesis, and activated dermal fibroblasts. The effects of the immunosuppressive agent bucillamine (SA 96) on fibroblasts and angiogenic factors have not been examined. SA 96, and particularly its metabolite SA 981, increased the levels of vascular endothelial growth factor (VEGF) mRNA and protein dose-dependently in dermal fibroblasts from patients with SSc and healthy control subjects without influencing cell viability. SSc fibroblast cultures showed consistently a higher inducibility of VEGF than cultures from healthy control subjects. Preincubation with the SP-1 inhibitor mithramycin as well as blockade of nuclear factor (NF)-kappaB signaling with pyrrolidine dithiocarbamate treatment and IkappaB transfection reduced significantly the transcription of VEGF, indicating that both transcription factors contribute to the activation of VEGF by SA 981. Specific binding of NF-kappaB protein to its binding site after treatment with SA 981 was confirmed by electrophoretic mobility shift assay. In contrast, SA 981 did not influence the stability of VEGF mRNA as analyzed with actinomycin D assays. The study provides evidence for a role of NF-kappaB in the transcriptional regulation of the VEGF gene. SA 96 might have positive aspects on the impaired angiogenesis in patients with SSc.

Cells, Cultured↗
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↗
PubMed2003

Fatal outcome of constrictive pericarditis in rheumatoid arthritis.

This is a report of a 39-year-old patient diagnosed with seropositive rheumatoid arthritis at the age of 17. The patient died 2 years after the onset of extra-articular cardiac symptoms. This case demonstrates the devastating course of progressive constrictive pericarditis under sole medical therapy and emphasizes the importance of early radical pericardectomy to avoid progression of disease and secondary complications with fatal outcome. Further, we discuss risk factors, diagnostic caveats, diagnostic tests and therapy of hemodynamically relevant contrictive pericarditis.

Adult↗
PubMed2003

Albumin-based drug delivery as novel therapeutic approach for rheumatoid arthritis.

We reported recently that albumin is a suitable drug carrier for targeted delivery of methotrexate (MTX) to tumors. Due to pathophysiological conditions in neoplastic tissue, high amounts of albumin accumulate in tumors and are metabolized by malignant cells. MTX, covalently coupled to human serum albumin (MTX-HSA) for cancer treatment, is currently being evaluated in phase II clinical trials. Because synovium of patients with rheumatoid arthritis (RA) shares various features observed also in tumors, albumin-based drug targeting of inflamed joints might be an attractive therapeutic approach. Therefore, the pharmacokinetics of albumin and MTX in a mouse model of arthritis was examined. Additionally, uptake of albumin by synovial fibroblasts of RA patients and the efficacy of MTX and MTX-HSA in arthritic mice were studied. The results show that when compared with MTX, significantly higher amounts of albumin accumulate in inflamed paws, and significantly lower amounts of albumin are found in the liver and the kidneys. The protein is metabolized by human synovial fibroblasts in vitro and in vivo. MTX-HSA was significantly more effective in suppression of the onset of arthritis in mice than was MTX. In conclusion, albumin appears to be a suitable drug carrier in RA, most likely due to effects on synovial fibroblasts, which might increase therapeutic efficacy and reduce side effects of MTX.

Albumins↗