Biomedical subjects

James Gardner-Thorpe

Publications and source records attributed to James Gardner-Thorpe.

9 recordsLinked to original sources

PubMed2004

Fibronectin-induced COX-2 mediates MMP-2 expression and invasiveness of rhabdomyosarcoma.

Although accumulating evidence suggests the importance of cyclooxygenase-2 (COX-2) and prostaglandin E(2) (PGE(2)) in the pathogenesis of many cancers, the mechanism by which this enzyme and its metabolite promote cancer progression is unknown. In this study, we investigated the role of COX-2 in fibronectin-induced up-regulation of rhabdomyosarcoma matrix metalloproteinase (MMP)-2 activity and cellular invasiveness. We tested three human rhabdomyosarcoma cell lines: RMS559, RD, and SJRH30. Cell attachment to fibronectin up-regulated both COX-2 expression and PGE(2) production and concomitantly enhanced MMP-2 activity. Exogenous PGE(2) stimulated MMP-2 promoter activity, increased MMP-2 expression, and increased cellular invasiveness. Aspirin and rofecoxib (non-selective and selective COX-2 inhibitor, respectively) each abolished fibronectin-associated induction of MMP-2 and induced dose-dependent reductions in cellular invasiveness. These data implicated a role for inducible COX-2 and PGE(2) in the regulation of rhabdomyosarcoma cellular invasiveness and MMP-2 activity.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2004

Lymphangiogenesis in tissue-engineered small intestine.

BACKGROUND: Lymphangiogenesis, the formation of lymphatic vessels, has not been reported in engineered tissue. The purpose of this study was to characterize lymphangiogenesis in tissue-engineered small intestine. METHODS: Biodegradable polymer scaffolds seeded with intestinal organoid units were implanted into syngenic recipient rats. Twenty-three neointestinal grafts were harvested from adult rat recipients 1 to 8 weeks postimplantation. Cells expressing the lymphatic endothelial marker vascular endothelial growth factor receptor (VEGFR)-3 were detected immunohistochemically. The lymphangiogenic growth factor VEGF-C was quantified by enzyme-linked immunoadsorbent assay. RESULTS: Between the first and eighth weeks, neointestinal cysts increased in volume and mass. Muscular and mucosal layers increased in thickness and developed to resemble normal intestine histologically. The proportion of neointestinal VEGFR-3-positive cells increased and ultimately, tubular structures developed that resembled lymphatics architecturally, were distinct from CD34-positive blood vessels, and lacked luminal erythrocytes. CONCLUSION: Lymphangiogenesis occurs in tissue-engineered small intestine. This is the first demonstration of lymphatic vessels in an engineered tissue.

Animals↗

PubMed2003

Effect of GLP-2 on mucosal morphology and SGLT1 expression in tissue-engineered neointestine.

Using tissue-engineering techniques, we have developed a neointestine that regenerates the structural and dynamic features of native small intestine. In this study, we tested neointestinal responsiveness to glucagon-like peptide 2 (GLP-2). Neointestinal cysts were engineered by seeding biodegradable polymers with neonatal rat intestinal organoid units. The cysts were matured and anastomosed to the native jejunum of syngeneic adult recipients. Animals were treated with GLP-2 [Gly2] (twice daily, 1 microg/g body wt) or vehicle alone (control) for 10 days. Rats were then killed, and tissues were harvested for analysis. Na+-glucose cotransporter (SGLT1) mRNA expression was assessed with Northern blotting and in situ hybridization. SGLT1 protein was localized by using immunofluorescence. GLP-2 administration resulted in 1.8- and 1.7-fold increases (P < 0.05) in neointestinal villus height and crypt depth, respectively. GLP-2 administration also resulted in a 2.4-fold increase (P < 0.01) in neomucosal SGLT1 mRNA expression. SGLT1 mRNA expression was localized to enterocytes throughout the villi, and SGLT1 protein was localized to the brush border of enterocytes along the entire length of villi from the neointestine of GLP-2-treated animals. The response of tissue-engineered neointestine to exogenous GLP-2 includes mucosal growth and enhanced SGLT1 expression. Therefore, tissue-engineering principles may help in dissecting the regulatory mechanisms mediating complex processes in the intestinal epithelium.

Animals↗

PubMed2003

The efficacy of tyrosine kinase inhibitors on human pancreatic cancer cell lines.

We attempted to determine potential therapeutic targets in pancreatic cancer by performing microarray analysis and targeted chemotherapy on three human pancreatic cancer cell lines. We used a microarray to screen 847 genes involved in cytokine signaling, signal transduction, and transcription. Tyrosine kinases represented a common target driving proliferation among the three cell types. We tested the ability of Gleevec (STI-571), Lavendustin, Herbimycin, and Genistein to inhibit the proliferation of cells in culture as assessed by the MTT assay.Eighteen genes were found to be commonly expressed by the three cell lines. Of these, six (33%) included tyrosine phosphorylation signaling as part of the pathway. The most highly expressed common transcript was the EphB3 receptor, which is a tyrosine kinase. Herbimycin and Genistein were able to inhibit the proliferation of all three cell lines in a dose dependent manner, with a mean IC(50) of 1.71 microM and 223 microM, respectively; whereas Lavendustin and Gleevec were ineffective in the inhibition of proliferation. Transcriptional profiling yielded common targets and insights into the biology of cells in culture. Herbimycin- and Genistein-based kinase inhibitors may offer potential and should be tested in other in vivo models for their ability to inhibit the growth of pancreatic cancer.

Benzamides↗

PubMed2003

Inhibition of tyrosine kinase Src suppresses pancreatic cancer invasiveness.

BACKGROUND: Src is a 60-kDa tyrosine kinase that plays a critical role in signal transduction associated with cell-extracellular matrix interactions. We tested the hypothesis that Src inhibition might suppress pancreatic cancer cellular invasiveness. METHODS: We tested the effects of pyrazolopyrimidine (a Src kinase-specific inhibitor) on 3 human pancreatic cancer cell lines: BXPC-3, MIAPaCa-2, and PANC-1. Src expression was assayed with Western blotting. Pyrazolopyrimidine-mediated inhibition of Src phosphorylation was confirmed by immunoprecipitation. Matrix metalloproteinase (MMP) activities and cellular invasive potential were assessed by use of zymography and Boyden chamber assays, respectively. Cell growth was assessed with the MTT assay. RESULTS: Src was expressed in all 3 pancreatic cancer cell lines tested. Pyrazolopyrimidine completely suppressed Src phosphorylation, inhibited MMP2 (72kDa) and MMP9 (92kDa) activities by 40% to 34% (P <.05), and suppressed cellular invasiveness by more than 90% (P <.05) in all 3 cell lines. Pyrazolopyrimidine had variable effects on cell growth: 50% reduction (P <.05) in BXPC-3, 7% reduction (P >.05) in MIAPaCa-2, and 22% reduction (P <.05) in PANC-1. CONCLUSIONS: Inhibition of Src signaling results in a marked reduction of pancreatic cancer cellular invasiveness. Src may represent a novel therapeutic target for this deadly cancer.

Cell Division↗

PubMed2003

Angiogenesis in tissue-engineered small intestine.

Tissue-engineered intestine offers promise as a potential novel therapy for short bowel syndrome. In this study we characterized the microvasculature and angiogenic growth factor profile of the engineered intestine. Twenty-three tissue-engineered small intestinal grafts were harvested from Lewis rat recipients 1 to 8 weeks after implantation. Architectural similarity to native bowel obtained from juvenile rats was assessed with hematoxylin and eosin-stained sections. Capillary density, measured after immunohistochemical staining for CD34, was expressed as number of capillaries per 1000 nuclei. Vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) tissue levels were measured by ELISA and normalized to total protein. Over the 8-week period cysts increased in volume (0.5 cm(3) at week 1 versus 12.6 cm(3) at week 8) and mass (1.30 +/- 0.29 versus 9.74 +/- 0.3 g; mean +/- SEM). Muscular and mucosal layers increased in thickness, but capillary density remained constant (82.95 +/- 4.81 capillaries per 1000 nuclei). The VEGF level was significantly higher in juvenile rat bowel than in engineered cyst (147.6 +/- 23.9 versus 42.3 +/- 3.4 pg/mg; p < 0.001). Tissue bFGF levels were also higher (315 +/- 65.48 versus 162.3 +/- 15.09 pg/mg; p < 0.05). The mechanism driving angiogenesis differs in engineered intestine and in normal bowel. VEGF and bFGF delivery may prove useful for bioengineering of intestine.

Aging↗

PubMed2003

Autoantibody-mediated inhibition of pancreatic cancer cell growth in an athymic (nude) mouse model.

INTRODUCTION: Antiribosomal P autoantibodies are detectable in 12-16% of patients with systemic lupus erythematosus. AIM: To assess whether antiribosomal P autoantibodies could be useful as a novel form of immunotherapy for pancreatic cancer. METHODOLOGY: Three pancreatic cancer cell lines were incubated with antiribosomal P or normal human immunoglobulin. Viability was assayed with MTT, and apoptosis was detected by TUNEL. MIA PaCa-2 cells were injected into athymic mice. Animals were treated with intraperitoneal antibody or control immunoglobulin. Serum antibody levels were measured by ELISA. Tumor nodule size was measured weekly. Binding of the antibody to tumors was demonstrated with fluorescent microscopy. RESULTS: Antiribosomal P antibody inhibited pancreatic cancer cell proliferation up to 54.6% (p < 0.01) and was associated with a threefold increase in the rate of apoptosis (p < 0.05). Tumor volume after 4 weeks of treatment was 23.2 mm3, versus 141.5 mm3 for the control group (p < 0.05). Apoptosis rate within the nodules was increased twofold, to 11.4%, in comparison with control (p < 0.05). CONCLUSION: Antiribosomal P autoantibody at levels similar to those that can exist in SLE inhibits the growth of pancreatic cancer cells, in vitro and in vivo. The mechanism involves surface binding and apoptosis.

Animals↗

PubMed2002

Differential display of expressed genes in pancreatic cancer cells.

Activating K-ras mutations occur in 80-95% of pancreatic cancers. The purpose of this study was to conduct an open, panoramic survey of gene expression, using K-ras status as the axis over which to sub-classify pancreatic cancers. Differential display was used to contrast mRNA purified from exponentially growing PANC-1 and Capan-2 cells (mutated K-ras) with Hs766T and BxPC-3 cells (wild-type). Differences were confirmed by Northern analysis. Twenty-five transcripts were differentially expressed by a factor of two or more. Four transcripts were over-expressed and twelve were under-expressed in the mutants relative to the wild-types. The transcripts most strikingly over-expressed by the mutant cell lines were MARCKS, DKFZp547C244, and RPLP2. The transcripts over-expressed by the wild-types were CEACAM6, cDNA AK026924, and myosin light chain-6. Profiling of gene expression with respect to K-ras mutation status may lead to new insights into pancreatic cancer pathogenesis, as well as to the identification of novel therapeutic targets.

Antigens, CD↗

PubMedDate not supplied

Ribosomal protein P2: a potential molecular target for antisense therapy of human malignancies.

BACKGROUND: Ribosomal protein P2 is an important component of protein translation machinery. We hypothesized that antisense-mediated depletion may disrupt the proteome of cancer cells. This study includes experiments to ascertain whether this could be a useful approach for cancer therapies. MATERIALS AND METHODS: MIA PaCa-2 and BxPC-3 cells were transfected with P2-antisense oligonucleotide or controls. Growth was assayed using MTT. Protein P2 was measured using Western blotting. Proteomes were compared using two-dimensional electrophoresis and changes were characterized by mass spectrometry. A macroarray was used to identify cancers which may be vulnerable. RESULTS: Antisense-P2 reduced P2 levels by 63% (p < 0.05) and inhibited BxPC-3 growth to 65% of control (p < 0.05). Seventeen (5.4%) proteins changed on two-dimensional electrophoresis including Rho C, translationally-controlled tumor protein, vinculin, LDH, ribosomal protein L23a, F-actin capping protein and eIF-3. Breast cancer underexpressed P2 compared to normal tissue (p < 0.001). CONCLUSION: Antisense-P2 technology has potential to slow growth of cancer cells. This effect is mediated through multiple proteomic changes.

Base Sequence↗