Biomedical subjects

Steven W Warmann

Publications and source records attributed to Steven W Warmann.

12 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↗

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

Primary repair of esophageal atresia in extremely low birth weight infants: a single-center experience and review of the literature.

BACKGROUND: Advances in neonatal intensive care have led to an increased survival of very low birth weight (VLBW, <1,500 g) and extremely low birth weight infants (ELBW, <1,000 g). Several abnormalities may occur in these children, e.g. esophageal atresia (EA), imperforate anus or abdominal wall defects. Correction of EA is often performed as a staged procedure in this group of patients. OBJECTIVES: To evaluate the feasibility of a primary correction of EA in 4 ELBW and VLBW infants. METHODS: Between 2002 and 2004, 4 infants below 1,200 g were operated on in our institution with a diagnosis of EA with lower tracheoesophageal fistula. Birth weight ranged from 780 to 1,120 g (median: 920 g), gestational age from 28 to 30 weeks. Treatment included closure of the tracheoesophageal fistula and primary anastomosis of the esophagus in a one-step procedure. RESULTS: Primary correction of EA and fistula repair was feasible in all children. Initially, all children had a normal passage of the esophagus as observed in barium swallowing. One child suffering from a leakage of the anastomosis was managed conservatively. Another infant suffered from spontaneous small bowel perforation 6 days after surgery, which was treated by laparotomy. One child developed stenosis of the esophagus and required a single dilatation 14 months after initial treatment. In the 4th child, a type II cleft syndrome was subsequently diagnosed, requiring secondary cleft repair together with semifundoplication. This child eventually died from cytomegalovirus pneumonia. CONCLUSIONS: Primary repair of EA and closure of a tracheoesophageal fistula is technically feasible and offers a good treatment option for ELBW and VLBW infants. Staged repair can be avoided. Infants with cleft syndrome are still a diagnostic and therapeutic challenge.

Anastomosis, Surgical↗

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

Laparoscopic cystojejunostomy as a treatment option for pancreatic pseudocysts in children--a case report.

Pancreatic pseudocysts are a rare entity in children for which many approaches have been described. We report on the case of a 5-year-old boy with a pancreatic pseudocyst after blunt abdominal trauma. The patient's clinical and laboratory examination findings had also revealed an acute pancreatitis. His diagnostic workup included ultrasound examination and magnetic resonance cholangiopancreatography. Two large cysts were found at the tail of the pancreas. No injury of the pancreatic or bile duct was found. The child underwent successful laparoscopic cystojejunostomy. The patient was free of complaints after more than 2 years of follow-up. Laparoscopic cystojejunostomy in children with pancreatic pseudocysts may represent an alternative treatment option for large pancreatic pseudocysts.

Abdominal Injuries↗

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↗

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

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↗

PubMedDate not supplied

Effects of P-glycoprotein modulation on the chemotherapy of xenotransplanted human hepatoblastoma.

Multidrug resistance (MDR) contributes to limited treatment results in human hepatoblastoma (HB). The MDR1 gene and its product P-glycoprotein (P-gP) has been identified as important factor in this development. In other tumors, P-gP modulation leads to a restored chemosensitivity of the cells. The aim of this study was to analyze the P-gP-modulating effects of PSC 833, a cyclosporine derivate, and verapamil on the chemotherapy of HB in vivo. HB from 2 patients were transplanted subcutaneously into nude mice NMRI (nu/nu). Animals were divided into 7 groups: Group 1 (Control); Group 2 (CDDP); Group 3 (DOXO); Group 4 (DOXO + verapamil); Group 5 (DOXO + PSC 833); Group 6 (CDDP + verapamil); and Group 7 (CDDP + PSC 833). If DOXO was administered (regardless of the combination), the dose was two times 60 mg/m2. If CDDP was administered, the dose was two times 27 mg/m2. When the chemosensitizers were administered, the doses for PSC 833 and for verapamil were four times 5 mg/kg body-weight. In the combined treatment groups the chemosensitizers were given ten minutes prior to CDDP and DOXO. Tumor volume developments and a-fetoprotein (AFP) alterations were assessed. Relative expression levels of the MDR1 gene after treatment were determined using a semiquantitative rT-PCR approach. In a mixed HB, both chemosensitizers combined with DOXO or CDDP produced a significant reduction of tumor growth (p = .0001-.00063) and AFP levels (p = .0006-.0128) compared to tumors treated with DOXO or CDDP only. Treatment results were identical to those in a less differentiated pure embryonal HB, but only in one case (DOXO + PSC 833, p = .031) significant. The chemosensitizers had no influence on the MDR1 gene expression. MDR1 modulators improve the efficiency of DOXO and CDDP treatment in xenotransplanted HB. They do not induce a further increase of drug resistance in the tumors. The data provide evidence that chemosensitizers might improve treatment results in patients with advanced or relapsed HB.

ATP Binding Cassette Transporter, Subfamily B, Mem↗