Biomedical subjects

Jörg Fuchs

Publications and source records attributed to Jörg Fuchs.

At least 19 recordsLinked to original sources

PubMed2007

Effects of standard chemotherapy on tumor growth and regulation of multidrug resistance genes and proteins in childhood rhabdomyosarcoma.

The prognosis of rhabdomyosarcoma (RMS) in advanced stages is still sobering. Therapy is limited due to local tumor recurrence, development of metastases and multidrug resistance. The aim of this study was to investigate the development of multidrug resistance in cell lines and in xenografts of alveolar and embryonal RMS treated according to the German Soft Tissue Sarcoma Study (CWS). Alveolar and embryonal RMS cell lines were treated with Vincristine, Topotecan, Carboplatin, Actinomycin D, or Ifosfamide. Expression levels of resistance-associated genes were assessed using Real time-PCR. Nude mice (NMRI nu/nu, n = 10 per group) underwent xenotransplantation of human embryonal or alveolar RMS. Animals were treated with standard chemotherapeutic drugs Vincristine, Topotecan, Carboplatin, Actinomycin D, or Ifosfamide according to treatment schedules of the CWS-study. Tumor sizes were measured and relative tumor volumes were calculated. Animals were sacrificed after 20 days and standard histology, Real-time-PCR for MDR1-, MRP-, LRP- and MDM2-gene as well as immunohistochemistry for MDR1-, LRP-, and MRP-protein were performed. In the cell lines, an up-regulation of MDR-1 gene was found in alveolar rhabdomyosarcoma. In embryonal rhabdomyosarcoma, an up-regulation of LRP and MRP was found. Standard chemotherapy of alveolar rhabdomyosarcoma resulted in a significant reduction of tumor growth (P < 0.05) in all groups. In embryonal rhabdomyosarcoma strongest effects were found after treatment with Ifosfamide, Vincristine and Carboplatin (P < 0.05). RT-PCR revealed a MDR1-dependent mechanism in alveolar rhabdomyosarcoma. In embryonal rhabdomyosarcoma, MDR1 occurred to a lower degree. Immunohistochemistry revealed correlating expression levels of multidrug resistance-associated proteins. The use of established chemotherapy on human RMS in vivo had strong effects on xenografts compared to their controls. In all cases, there was only a reduction of tumor growth, but not a complete eradication of the tumors. Chemotherapy seemed to upregulate the expression of resistance-associated genes in vitro and in vivo. The mechanism of multidrug resistance depends on the tumor subtype. Therefore, further investigations will be required to evaluate multidrug resistance in patients and to investigate new modalities for a reversal of multidrug resistance.

Analysis of Variance↗

PubMed2007

Treatment of cystic nephroma and cystic partially differentiated nephroblastoma--a report from the SIOP/GPOH study group.

PURPOSE: Cystic partially differentiated nephroblastoma is a rare variant of Wilms tumor, and might be confused with cystic nephroma. Definitive diagnosis can only be made by histological examination. Therefore, initiation of therapy, either primary nephrectomy or preoperative chemotherapy, might create a dilemma when radiological diagnosis is doubtful. MATERIALS AND METHODS: To define treatment strategies for these entities, we reviewed the records of 1,245 patients enrolled in SIOP (International Society of Pediatric Oncology) trials 93-01 and 2001 GPOH (German Society of Pediatric Oncology and Hematology) between July 1993 and August 2004. Data were collected retrospectively. Therapy, outcome and preoperative management were evaluated. To confirm diagnosis of cystic nephroma/partially differentiated nephroblastoma, all patients underwent review by the Reference Pathology Center of SIOP/GPOH. RESULTS: A total of 14 patients with diagnoses of cystic nephroma (7) and cystic partially differentiated nephroblastoma (7) were identified. Median patient age at diagnosis was 1 year (0.46 to 3). Two patients received preoperative chemotherapy. Primary nephrectomy was performed in 12 patients. Two patients underwent partial nephrectomy. In 1 child postoperative chemotherapy was administered. None of the patients had progression of disease or recurrence. Overall survival was 100%. Median followup was 2.41 years (0.3 to 9). CONCLUSIONS: In cystic renal tumors radiological findings should always be reviewed by the reference radiologist of the treatment protocol study group. Irrespective of the chosen therapy, outcome of cystic nephroma and cystic partially differentiated nephroblastoma is favorable. Even in large international trials the number of patients with cystic nephroma or cystic partially differentiated nephroblastoma is too small for statistical analysis.

Child, Preschool↗

PubMed2006

Optimizing vector application for gene transfer into human hepatoblastoma cells.

Gene targeting is currently of distinct interest as an innovative additive treatment option in various malignancies. Its role in pediatric liver tumors has not yet been evaluated thoroughly. For the first time the authors systematically analyzed both lipid-based transfection as well as transduction with adenovirus vectors (Ad) and Sendai virus vectors (SeVV) in order to optimize gene transfer into hepatoblastoma (HB) cells. Two HB cell lines were infected with Ad or SeVV coding for green fluorescent protein (Ad-GFP, SeVV-GFP); transduction efficiencies and apoptosis were assessed using flow cytometry. Furthermore, lipofection of HB cell lines with plasmid-constructs comprising liver-specific promoters was performed using Lipofectamine 2000 and FuGENE 6; lipofection efficiency was monitored by flow cytometry, microscopy, and luciferase activity. The Ad-GFP showed higher transduction rates (61-86%) than the SeVV-GFP (4-24%) depending on the HB cell line used. Infections with first generation SeVV vectors (SeVV-GFP) led to increased target cell apoptosis (7-43%) compared to Ad-GFP (4-16%). The Lipofectamine 2000 revealed a higher transfection efficiency than the FuGENE 6 for both HB cell lines tested. The liver-specific promoters were found to be differently active in the HB cell lines. This study delineates recombinant adenovirus vectors as a promising tool for gene transduction in the HB cells. Furthermore, enhanced activity of the liver-specific promoters in HUH6 cells compared to HepT1 cells supports the observation of varying biological behavior in histologically differing HB tissues.

Adenoviridae↗

PubMed2006

Chromosomal histone modification patterns--from conservation to diversity.

The organization of DNA into chromatin regulates expression and maintenance (replication, repair, recombination, segregation) of genetic information in a dynamic manner. The N-terminal tails of the nucleosomal core histones are subjected to post-translational modifications such as acetylation, methylation, phosphorylation, ubiquitination, glycosylation, ADP-ribosylation, carbonylation and sumoylation. These modifications, together with DNA methylation, control the folding of the nucleosomal array into higher order structures and mediate signalling for cellular processes. Although histones and their modifications are highly conserved, recent data show that chromosomal distribution of individual modifications (acetylation, methylation, phosphorylation) can differ along the cell cycle as well as among and between groups of eukaryotes. This implies the possibility of evolutionary divergence in reading the "histone code".

Acetylation↗

PubMed2006

In vitro gene targeting in human hepatoblastoma.

Poor treatment results in advanced hepatoblastoma (HB) made alternative treatment approaches desirable. Gene-directed tumor therapy is increasingly investigated in different malignancies. The aim of this study was to analyze possible alternatives of gene transfer into HB cells and to study therapeutic applications based on different strategies. Liposomal transfection of HB cells was assessed using liver-specific promoters, and adenovirus and Sendai virus transductions were performed in vitro. Transfer efficiencies were measured via flow cytometry determining expression of vector-encoded marker gene green fluorescent protein. Gene silencing of the anti-apoptotic bcl-2 gene in HUH6 cells was performed using lipofection of small interfering RNA (siRNA). Additionally, suicide gene therapy was carried out through a yeast-derived cytosine deaminase (YCD)-combined yeast uracil phosphoribosyltransferase (YUPRT)-based adenovirus-mediated gene transfer, leading to a potent intracellular prodrug transformation of 5-fluorocytosine into 5-fluorouracil. Treatment efficiencies were monitored via MTT viability assay. Highest gene transfer rates (86%) were observed using adenovirus transduction. We furthermore observed a significant therapeutic effect of adenovirus-mediated YCD::YUPRT suicide gene transfer. Liposomal-mediated anti-bcl-2 siRNA transfer led to a significant improvement of cisplatin treatment in HUH6 cells. Liver-specific promoters were found to be strongly active in HUH6 cells (mixed HB-derived), but less active in HepT1 cells (embryonal HB-derived). Liposomal transfection and viral transduction are effective approaches to genetically manipulate HB cells in vitro. For the first time, we demonstrate a positive effect of siRNA gene silencing in this malignancy. Additionally, we successfully investigated a model of adenovirus-based suicide gene therapy in HB cell cultures. Our data strongly encourage further studies assessing these alternative treatment approaches.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Visualization of xenotransplanted human rhabdomyosarcoma after transfection with red fluorescent protein.

BACKGROUND/AIMS: Discosoma sp red fluorescent protein (DsRed2) is a newly developed marker for in vivo labeling studies in different biologic systems. After vector transfection, DsRed2 is expressed in mammalian cells and can be detected by fluorescence microscopy. The aims of this study were to establish a DsRed2-transfected human rhabdomyosarcoma (RMS) cell line and to perform a xenotransplantation on nude mice to use imaging as a tool for further basic research studies on this neoplasm. PROCEDURE: The human alveolar RMS cell line Rh30 was transfected with the pDsRed2-N1 vector by lipofection. The DsRed2-positive cells were sorted out by fluorescence-activated cell sorting analysis 96 hours after transfection and selected in culture with G418. Expression of DsRed2 messenger RNA was assessed using single-cell reverse transcriptase polymerase chain reaction after laser microdissection. Transfected and parental cells were characterized cytologically, cytogenetically, immunohistochemically, and in vivo after subcutaneous injection in NMRI (nu/nu) nude mice. RESULTS: After vector transfection, a pure and stable DsRed2-positive cell line was established by monoclonal growth of the cells. Reverse transcriptase polymerase chain reaction revealed constant expression of DsRed2 messenger RNA in fluorescencing cells. There was no difference between transfected and parental cells by means of cell morphology and desmin expression. Clonal cells (1 x 10(6)) were used for xenotransplantation. Tumors were visualized noninvasively through the skin of the mice using specific emission and excitation filters. Tumor vascularization and vessel growth could be discriminated from tumor tissue using this imaging system. CONCLUSION: This is the first report on successful transfection of an RMS cell line with red fluorescent protein followed by xenotransplantation into nude mice. This model can serve as an imaging tool for in vivo studies investigating tumor biology and metastases of human RMS.

Animals↗

PubMed2006

Giant mullerian duct cyst with malignant transformation in 15-year-old boy.

We present a case of a giant mullerian duct cyst with components of clear cell adenocarcinoma in a 15-year-old boy. Complete tumor resection was achieved in two steps. The presented case with a combination that has never been described before may change the perspective of treatment approaches for this rare entity.

Adenocarcinoma, Clear Cell↗

PubMed2006

Chromosome arrangement and nuclear architecture but not centromeric sequences are conserved between Arabidopsis thaliana and Arabidopsis lyrata.

In contrast to the situation described for mammals and Drosophila, chromosome territory (CT) arrangement and somatic homologous pairing in interphase nuclei of Arabidopsis thaliana (n = 5) are predominantly random except for a more frequent association of the chromosomes bearing a homologous nucleolus organizer region. To find out whether this chromosome arrangement is also characteristic for other species of the genus Arabidopsis, we investigated Arabidopsis lyrata ssp. lyrata (n = 8), one of the closest relatives of A. thaliana. First, we determined the size of each chromosome and chromosome arm, the sequence type of centromeric repeats and their distribution between individual centromeres and the position of the 5S/45S rDNA arrays in A. lyrata. Then we demonstrated that CT arrangement, homologous pairing and sister chromatid alignment of distinct euchromatic and/or heterochromatic regions within A. lyrata interphase nuclei are similar to that in A. thaliana nuclei. Thus, the arrangement of interphase chromosomes appears to be conserved between both taxa that diverged about 5 million years ago. Since the chromosomes of A. lyrata resemble those of the presumed ancestral karyotype, a similar arrangement of interphase chromosomes is also to be expected for other closely related diploid species of the Brassicaceae family.

Arabidopsis↗

PubMed2005

A complicated case of bilateral cystic nephroma in a 16-month-old boy.

We describe a case of a progressive bilateral cystic nephroma (BCN) in a child undergoing a multistaged surgical procedure. After partial resection of the left sided tumor, a progress occurred on that side and the left kidney had to be removed 10 weeks later. After 35 months a tumor progression was observed on the right side together with an ureteral obstruction leading to a decreased renal function. In a third operation a complete tumor resection on the right side was achieved through longitudinal partial nephrectomy, reconstruction of the renal pelvis, and reanastomosis of the reconstructed pelvis and ureter. The patient showed no evidence of the disease at 28 months of follow-up. The presented case provides an evidence that in BCN a tumor progress may occur after multistaged surgical approaches. A single-staged complete tumor resection with renal salvage techniques seems indicated.

Disease Progression↗

PubMed2005

Electromagnetic fields affect transcript levels of apoptosis-related genes in embryonic stem cell-derived neural progenitor cells.

Mouse embryonic stem (ES) cells were used as an experimental model to study the effects of electromagnetic fields (EMF). ES-derived nestin-positive neural progenitor cells were exposed to extremely low frequency EMF simulating power line magnetic fields at 50 Hz (ELF-EMF) and to radiofrequency EMF simulating the Global System for Mobile Communication (GSM) signals at 1.71 GHz (RF-EMF). Following EMF exposure, cells were analyzed for transcript levels of cell cycle regulatory, apoptosis-related, and neural-specific genes and proteins; changes in proliferation; apoptosis; and cytogenetic effects. Quantitative RT-PCR analysis revealed that ELF-EMF exposure to ES-derived neural cells significantly affected transcript levels of the apoptosis-related bcl-2, bax, and cell cycle regulatory "growth arrest DNA damage inducible" GADD45 genes, whereas mRNA levels of neural-specific genes were not affected. RF-EMF exposure of neural progenitor cells resulted in down-regulation of neural-specific Nurr1 and in up-regulation of bax and GADD45 mRNA levels. Short-term RF-EMF exposure for 6 h, but not for 48 h, resulted in a low and transient increase of DNA double-strand breaks. No effects of ELF- and RF-EMF on mitochondrial function, nuclear apoptosis, cell proliferation, and chromosomal alterations were observed. We may conclude that EMF exposure of ES-derived neural progenitor cells transiently affects the transcript level of genes related to apoptosis and cell cycle control. However, these responses are not associated with detectable changes of cell physiology, suggesting compensatory mechanisms at the translational and posttranslational level.

Apoptosis↗

PubMed2005

Extracellular calcium deficiency and ryanodine inhibit Eimeria tenella sporozoite invasion in vitro.

An in vitro assay system with Eimeria tenella sporozoites was used to study the effects of extracellular calcium and active agents affecting the invasion of parasites into host cells. At concentrations of 900 microM Ca(2+) and less the invasion rates were distinctly decreased. Ryanodine, a herbal alkaloid known for binding to internal Ca(2+) channels (ryanodine receptors), showed an inhibitory effect on E. tenella sporozoite invasion. Preincubation tests and staining with a fluorescent derivative of ryanodine assured that the compound bound specifically to the sporozoites and affected them rather than the host cells.

Animals↗

PubMed2005

Three-dimensional visualization and virtual simulation of resections in pediatric solid tumors.

PURPOSE: Three-dimensional visualization of solid tumors is possible because of high-resolution computed tomography and magnetic resonance imaging scans. However, additional preoperative information is often desirable in complex malignancies. For the first time, the authors present a model of preoperative 3-dimensional visualization and virtual resections in pediatric solid tumors. METHODS: Image analysis of various pediatric tumors was performed using the research software HepaVision2 (MeVis, Bremen). Organs, tumors, and the vascular system were extracted from multislice computed tomography scans. After hierarchical analysis of the vascular system, territories supplied or drained by the major vascular branches were calculated. Results were explored and virtual resections of organs were carried out using the research software InterventionPlanner (MeVis, Bremen). Data were correlated to intraoperative findings. RESULTS: Four hepatic malignancies, 4 renal tumors, and 3 other neoplasms were analyzed. The technique of 3-dimensional visualization was feasible for all investigated children (mean age 5 years and 9 months). Spatial relations between physiological and pathological structures were identified, and anatomical structures (vessels, tumor tissue, and organ parenchyma) were determined using colorimetric encoding. Virtual simulations of tumor resection were used successfully for planning of surgical procedures in the hepatic and renal tumors. CONCLUSIONS: The technique of 3-dimensional tumor visualization and virtual simulation of tumor resections provides the basis for a successful planning of complex tumor resections in children. The efficiency of these techniques should be further analyzed in series with higher numbers and differentiations of tumors.

Child↗

PubMed2005

New trends in tumor biology: transfection of a human hepatoblastoma cell line with green fluorescent protein.

BACKGROUND/PURPOSE: Enhanced green fluorescent protein (eGFP) is widely used as a marker in different biologic systems. After vector transfection, eGFP is expressed by eukaryotic cells and can be visualized using fluorescent microscopy. The aim of this study was to establish an eGFP-transfected human hepatoblastoma (HB) cell line as tool for further basic research studies. METHODS: The HB cell line HUH6 was transfected with the pEGFP-N1 vector by liposomal transfection. Enhanced green fluorescent protein-positive cells were sorted out by fluorescence-activated cell sort and selected using G418 resistance. Expression of eGFP-messenger RNA was assessed by single-cell reverse transcriptase polymerase chain reaction after laser microdissection. Original and transfected cells were compared biologically and cytomorphologically. RESULTS: Vector transfection produced up to 15% eGFP-positive cells. After fluorescence-activated cell sort and G418 selection, a pure cell line was established with 100% eGFP-positive cells. Reverse transcriptase polymerase chain reaction revealed constant expression of eGFP-messenger RNA in fluorescending cells. Analysis of cell characteristics revealed no differences between transfected and original cells. CONCLUSIONS: For the first time, the authors established an eGFP-transfected HB cell line. This cell line can serve as a promising tool for further studies investigating HB in vitro and in vivo. Our model might also be a basis for similar work on other pediatric solid tumors.

Flow Cytometry↗

PubMed2005

Protective effect of tumor necrosis factor alpha antibody on experimental necrotizing enterocolitis in the rat.

BACKGROUND: Necrotizing enterocolitis (NEC) is a common and devastating disorder of premature infants. Elevated proinflammatory cytokines, especially tumor necrosis factor alpha (TNF-alpha), have been implicated in the pathogenesis of NEC. The aim of this study was to evaluate the effects of TNF-alpha on the inflammatory response in NEC by immunoneutralizing TNF-alpha with a selective antibody. METHODS: Neonatal Sprague-Dawley rats were divided in 3 groups: group 1 (n = 20), a NEC-like enterocolitis was induced by formula feeding, asphyxia, and cold exposure; group 2 (n = 9), animals were treated like in group 1 and additionally received TNF-alpha antibody intraperitoneally; and group 3 (n = 17), animals were dam-fed (controls). Animals were killed in case of imminent death or after 96 hours. Specimens from small bowel were processed for blinded histologic (H&E) and immunhistologic (myeloperoxidase [MPO]) analysis. RESULTS: In group 1, animals developed severe NEC (mean NEC score, 3.28 +/- 0.32; mean MPO, 65.85 +/- 9.46). In group 2, animals developed mild NEC (mean NEC score, 1.72 +/- 0.41; mean MPO, 34.33 +/- 9.69; P < .05). In group 3, no NEC was induced (mean NEC score, 0.0 +/- 0; mean MPO, 6 +/- 1.32; P < .05). CONCLUSION: Tumor necrosis factor alpha antibody may have an attenuating effect on experimental NEC in rats.

Animals↗

PubMed2004

Chromosome territory arrangement and homologous pairing in nuclei of Arabidopsis thaliana are predominantly random except for NOR-bearing chromosomes.

Differential painting of all five chromosome pairs of Arabidopsis thaliana revealed for the first time the interphase chromosome arrangement in a euploid plant. Side-by-side arrangement of heterologous chromosome territories and homologous association of chromosomes 1, 3 and 5 (on average in 35-50% of nuclei) are in accordance with the random frequency predicted by computer simulations. Only the nucleolus organizing region (NOR)-bearing chromosome 2 and 4 homologs associate more often than randomly, since NORs mostly attach to a single nucleolus. Somatic pairing of homologous approximately 100 kb segments occurs less frequently than homolog association, not significantly more often than expected at random and not simultaneously along the homologs. Thus, chromosome arrangement in Arabidopsis differs from that in Drosophila (characterized by somatic pairing of homologs), in spite of similar genome size, sequence organization and chromosome number. Nevertheless, in up to 31.5% of investigated Arabidopsis nuclei allelic sequences may share positions close enough for homologous recombination.

Animals↗

PubMed2004

Dual histone H3 methylation marks at lysines 9 and 27 required for interaction with CHROMOMETHYLASE3.

Both DNA methylation and post-translational histone modifications contribute to gene silencing, but the mechanistic relationship between these epigenetic marks is unclear. Mutations in two Arabidopsis genes, the KRYPTONITE (KYP) histone H3 lysine 9 (H3K9) methyltransferase and the CHROMOMETHYLASE3 (CMT3) DNA methyltransferase, cause a reduction of CNG DNA methylation, suggesting that H3K9 methylation controls CNG DNA methylation. Here we show that the chromodomain of CMT3 can directly interact with the N-terminal tail of histone H3, but only when it is simultaneously methylated at both the H3K9 and H3K27 positions. Furthermore, using chromatin immunoprecipitation analysis and immunohistolocalization experiments, we found that H3K27 methylation colocalizes with H3K9 methylation at CMT3-controlled loci. The H3K27 methylation present at heterochromatin was not affected by mutations in KYP or in several Arabidopsis PcG related genes including the Enhancer of Zeste homologs, suggesting that a novel pathway controls heterochromatic H3K27 methylation. Our results suggest a model in which H3K9 methylation by KYP, and H3K27 methylation by an unknown enzyme provide a combinatorial histone code for the recruitment of CMT3 to silent loci.

Amino Acid Sequence↗

PubMed2004

Behaviour of nucleolus organizing regions (NORs) and nucleoli during mitotic and meiotic divisions in budding yeast.

Spatial organization and segregation behaviour in mitosis and meiosis of nucleoli and NORs of Saccharomyces cerevisiae were examined by fluorescence in situ hybridization (FISH) and immunostaining, and integrated with previous ultrastructural studies. Our observations suggest that in interphase the NOR-bearing chromosome arm forms a hairpin-shaped loop inside the nucleolus, unlike the other arms which adopt a Rabl-like orientation. Prior to mitosis and meiosis, the appearance of the NOR changes from puffed to thread-shaped. In mitosis, it is stretched between the mother and daughter nuclei and seems to be among the last regions where chromatids separate. The nucleoli remain intact and split at the end of anaphase. Similarly, during meiosis I, intact nucleoli trail behind the separating homologous NORs and are partitioned equally to the two half-nuclei. During the second meiotic division, however, the nucleolus, together with a major portion of the nucleoplasm and the nuclear pore complexes, are not included in the spores. The behaviour of nucleoli in meiotic mutants and a strain with extrachromosomal rDNA suggests that they are not actively extruded but rather are lost due to their detachment from the separating chromosomes. We discuss the possibility that the exclusion of the nucleolus from the spores serves the disposal of agents that resort to the nucleolus, and has a role in spore dormancy or rejuvenation.

Cell Nucleolus↗

PubMed2003

A prospective study of minimally invasive techniques in pediatric surgical oncology: preliminary report.

BACKGROUND: Thoracoscopic and laparoscopic techniques play a major role in pediatric surgery. However, minimally invasive surgery (MIS) has not yet established itself in pediatric surgical oncology. The authors present a prospective study investigating the role of MIS in children with cancer. PROCEDURE: All children with abdominal or thoracic tumors requiring surgery were registered between September, 2000 and February, 2002. Decisions regarding procedures and approaches-conventional or minimally invasive-were made by the interdisciplinary team. Data on diagnoses, surgical procedures, complications, and conversion rates were registered prospectively. RESULTS: Seventy-four patients received 78 operations, 21 (26.9%) of the 78 operations were minimally invasive. Seven of 16 tumor biopsies (43.8%) and 9 of 57 tumor resections (15.8%) were performed using MIS, which was also exclusively used for diagnostic interventions. Conversions to standard techniques only occurred in 5 of 9 tumor resections. No major complications were encountered in the MIS group. CONCLUSIONS: MIS was practical in every fourth patient in our experience so far. It proved to be an excellent approach in diagnostic interventions and tumor biopsies, whereas efficacy is limited in tumor resections. Further factors (tumor recurrence, trocar site recurrence, tumor growth, and dissemination after CO(2) insufflation) have to be evaluated. Our data encourage the continuation of the study.

Abdominal Neoplasms↗