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ABSTRACT: Sera mass spectrometry (MS) peak differences were analyzed from 35 ovarian cancer patients and 16 disease-free individuals. "Leave one out" cross validation was used to assign "% cancer peaks" in control and ovarian cancer sera samples. Sera MS discriminated stage I/II and stage III/V ovarian cancer patients versus controls with ROC curve area values of 0.82 and 0.92. Test sensitivities for ovarian cancer stage I/II and III/V were 80% and 93% respectively. These results indicate that MS is useful for distinguishing sera from early-stage ovarian cancer patients, and has potential as a test for early detection of this disease.
Cancer Investigation 12/2011; 30(2):189-97. · 1.85 Impact Factor
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ABSTRACT: Goals of this study were to analyze the ability of mass spectrometry serum profiling to distinguish non-small cell lung adenocarcinoma from squamous cell carcinoma patients and healthy controls. Sera were obtained from 19 adenocarcinoma patients, 24 squamous cell carcinoma patients, and 21 controls. Identifications of significant mass-to-charge ratio (m/z) peak differences between these groups were performed using t-tests. A "leave one out" cross-validation procedure yielded discriminatory lung adenocarcinoma versus squamous cell carcinoma p and ROC curve values of <.0001 and 0.92, respectively. Test sensitivity and specificity were 84% and 79%, respectively. This approach could aid in lung cancer diagnosis and sub-typing.
Cancer Investigation 12/2011; 30(2):180-8. · 1.85 Impact Factor
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James R Hocker,
Marvin D Peyton, Megan R Lerner,
Stephanie L Mitchell,
Stan A Lightfoot,
Theresa J Lander,
Leah M Bates-Albers,
Nicole T Vu,
Rushie J Hanas,
Thomas C Kupiec,
Daniel J Brackett,
Jay S Hanas
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ABSTRACT: The goal of this study was to evaluate the usefulness of electrospray ionization-mass spectrometry (ESI-MS) technology to distinguish sera of early-stage lung cancer patients from control individuals. ESI-MS m/z (mass divided by charge) data were generated from sera of 43 non-small cell lung cancer patients (pathological stages I and II) and 21 control individuals. Identifications of m/z peak area significances between cancer and control ESI-MS sera spectra were performed using t-tests. A "leave one out" cross validation procedure, which mimics blinded sera analysis and corrects for "over-fitting" of data, yielded discriminatory cancer versus control distribution p value and ROC curve area value of <0.001 and 0.87, respectively. Analysis without the "leave one out" cross validation procedure yielded a ROC curve area of 0.99 for discrimination of sera from lung cancer patients versus control individuals. Predictive value measurements revealed overall test efficiency and sensitivity for distinguishing sera from lung cancer patients from controls (using "leave one out" cross validation) of 80% and 84%, respectively. ESI-MS serum analysis between control individuals and lung cancer patients who smoked or did not smoke had p values in ranges indicating that smoking effects are not pronounced in our analysis. These studies indicate that ESI-MS analyses of sera from early stage non-small cell lung cancer patients were helpful in distinguishing these patients from control individuals.
Lung cancer (Amsterdam, Netherlands) 05/2011; 74(2):206-11. · 3.14 Impact Factor
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James R Hocker, Megan R Lerner,
Stephanie L Mitchell,
Stan A Lightfoot,
Theresa J Lander,
Aurelien A Quillet,
Rushie J Hanas,
Marvin D Peyton,
Russell G Postier,
Daniel J Brackett,
Jay S Hanas
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ABSTRACT: This study evaluated the usefulness of electrospray mass spectrometry to distinguish sera of early-stage pancreatic cancer patients from disease-free individuals. Sera peak data were generated from 33 pancreatic cancer patients and 30 disease-free individuals. A "leave one out" cross-validation procedure discriminated stage I/II pancreatic cancer versus disease-free sera with a p value <.001 and a receiver-operator characteristic curve area value of 0.85. Predictive values for cancer stage I/II test efficiency, specificity, and sensitivity were 78%, 77%, and 79%, respectively. These studies indicate that electrospray mass spectrometry is useful for distinguishing sera of early-stage pancreatic cancer patients from disease-free individuals.
Cancer Investigation 02/2011; 29(2):173-9. · 1.85 Impact Factor
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ABSTRACT: Numerous microRNAs (miRNAs) are reported as differentially expressed in cancer, however the consequence of miRNA deregulation in cancer is unknown for many miRNAs. We report that two miRNAs located on chromosome 17p13, miR-132 and miR-212, are over-expressed in pancreatic adenocarcinoma (PDAC) tissues. Both miRNAs are predicted to target the retinoblastoma tumor suppressor, Rb1. Validation of this interaction was confirmed by luciferase reporter assay and western blot in a pancreatic cancer cell line transfected with pre-miR-212 and pre-miR-132 oligos. Cell proliferation was enhanced in Panc-1 cells transfected with pre-miR-132/-212 oligos. Conversely, antisense oligos to miR-132/-212 reduced cell proliferation and caused a G(2)/M cell cycle arrest. The mRNA of a number of E2F transcriptional targets were increased in cells over expressing miR-132/-212. Exposing Panc-1 cells to the β2 adrenergic receptor agonist, terbutaline, increased the miR-132 and miR-212 expression by 2- to 4-fold. We report that over-expression of miR-132 and miR-212 result in reduced pRb protein in pancreatic cancer cells and that the increase in cell proliferation from over-expression of these miRNAs is likely due to increased expression of several E2F target genes. The β2 adrenergic pathway may play an important role in this novel mechanism.
Biochemical and Biophysical Research Communications 02/2011; 406(4):518-23. · 2.48 Impact Factor
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Sripathi M Sureban,
Randal May,
Stan A Lightfoot,
Aimee B Hoskins, Megan Lerner,
Daniel J Brackett,
Russell G Postier,
Rama Ramanujam,
Altaf Mohammed,
Chinthalapally V Rao,
James H Wyche,
Shrikant Anant,
Courtney W Houchen
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ABSTRACT: Pancreatic cancer is an exceptionally aggressive disease in great need of more effective therapeutic options. Epithelial-mesenchymal transition (EMT) plays a key role in cancer invasion and metastasis, and there is a gain of stem cell properties during EMT. Here we report increased expression of the putative pancreatic stem cell marker DCAMKL-1 in an established KRAS transgenic mouse model of pancreatic cancer and in human pancreatic adenocarcinoma. Colocalization of DCAMKL-1 with vimentin, a marker of mesenchymal lineage, along with 14-3-3 σ was observed within premalignant PanIN lesions that arise in the mouse model. siRNA-mediated knockdown of DCAMKL-1 in human pancreatic cancer cells induced microRNA miR-200a, an EMT inhibitor, along with downregulation of EMT-associated transcription factors ZEB1, ZEB2, Snail, Slug, and Twist. Furthermore, DCAMKL-1 knockdown resulted in downregulation of c-Myc and KRAS through a let-7a microRNA-dependent mechanism, and downregulation of Notch-1 through a miR-144 microRNA-dependent mechanism. These findings illustrate direct regulatory links between DCAMKL-1, microRNAs, and EMT in pancreatic cancer. Moreover, they demonstrate a functional role for DCAMKL-1 in pancreatic cancer. Together, our results rationalize DCAMKL-1 as a therapeutic target for eradicating pancreatic cancers.
Cancer Research 02/2011; 71(6):2328-38. · 7.86 Impact Factor
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ABSTRACT: Limited information is available regarding systemic changes in mammals associated with exposures to petroleum/hydrocarbon fuels. In this study, systemic toxicity of JP-8 jet fuel was observed in a rat inhalation model at different JP-8 fuel vapor concentrations (250, 500, or 1000 mg/m(3), for 91 days). Gel electrophoresis and mass spectrometry sequencing identified the alpha-2 microglobulin protein to be elevated in rat kidney in a JP-8 dose-dependent manner. Western blot analysis of kidney and lung tissue extracts revealed JP-8 dependent elevation of inducible heat shock protein 70 (HSP70). Tissue changes were observed histologically (hematoxylin and eosin staining) in liver, kidney, lung, bone marrow, and heart, and more prevalently at medium or high JP-8 vapor phase exposures (500-1000 mg/m(3)) than at low vapor phase exposure (250 mg/m(3)) or non-JP-8 controls. JP-8 fuel-induced liver alterations included dilated sinusoids, cytoplasmic clumping, and fat cell deposition. Changes to the kidneys included reduced numbers of nuclei, and cytoplasmic dumping in the lumen of proximal convoluted tubules. JP-8 dependent lung alterations were edema and dilated alveolar capillaries, which allowed clumping of red blood cells (RBCs). Changes in the bone marrow in response to JP-8 included reduction of fat cells and fat globules, and cellular proliferation (RBCs, white blood cells-WBCs, and megakaryocytes). Heart tissue from JP-8 exposed animals contained increased numbers of inflammatory and fibroblast cells, as well as myofibril scarring. cDNA array analysis of heart tissue revealed a JP-8 dependent increase in atrial natriuretic peptide precursor mRNA and a decrease in voltage-gated potassium (K+) ion channel mRNA.
Toxicology mechanisms and methods 03/2010; 20(4):204-12. · 1.03 Impact Factor
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ABSTRACT: Autophagy is the regulated process cells use to recycle nonessential, redundant, or inefficient components and is an adaptive response during times of stress. In addition to its function in enabling the cell to gain vital nutrients in times of stress, autophagy can also be involved in elimination of intracellular microorganisms, tumor suppression, and antigen presentation. Because of difficulty in diagnosing autophagy, few clinical studies have been performed. This study examined whether autophagy occurs in hepatocytes during sepsis. Electron microscopy (EM) was performed on liver samples obtained from both an observational clinical cohort of six septic patients and four control patients as well as liver specimens from mice with surgical sepsis (by cecal ligation and puncture) or sham operation. EM demonstrated increased autophagic vacuoles in septic vs nonseptic patients. Randomly selected fields (3000 microm(2)) from control and septic patients contained 1.2+/-1.5 vs 5.3+/-3.3 (mean+/-s.d.) complex lysosomal/autophagolysosomal structures per image respectively (P<0.001). In rare instances, hepatocytes with autophagic vacuoles appeared to be unequivocally committed to death. Membrane alterations (membrane vacuoles, invagination into adjacent organelles, and myelin figure-like changes) occur in a subpopulation of mitochondria in sepsis, but other hepatocyte organelles showed no consistent ultrastructural injury. Findings in murine sepsis paralleled those of patients, with 7.2+/-1.9 vs 38.7+/-3.9 lysosomal/autophagolysosomal structures in sham and septic mice, respectively (P=0.002). Quantitative RT-PCR demonstrated that sepsis induced the upregulation of select apoptosis and cytokine gene expression with minimal changes in the core autophagy genes in liver. In conclusion, hepatocyte autophagic vacuolization increases during sepsis and is associated with mitochondrial injury. However, it is not possible to determine whether the increase in autophagic vacuolization is an adaptive response or a harbinger of cell death.
Laboratory Investigation 02/2009; 89(5):549-61. · 3.64 Impact Factor
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ABSTRACT: Enterococcus faecalis is an intestinal commensal that cannot synthesize porphyrins and only expresses a functional respiratory chain when provided with exogenous haematin. In the absence of haematin, E. faecalis reverts to fermentative metabolism and produces extracellular superoxide that can damage epithelial-cell DNA. The acute response of the colonic mucosa to haematin-starved E. faecalis was identified by gene array. E. faecalis was inoculated into murine colons using a surgical ligation model that preserved tissue architecture and homeostasis. The mucosa was exposed to haematin-starved E. faecalis and compared with a control consisting of the same strain grown with haematin. At 1 h post-inoculation, 6 mucosal genes were differentially regulated and this increased to 42 genes at 6 h. At 6 h, a highly significant biological interaction network was identified with functions that included nuclear factor-kappaB (NF-kappaB) signalling, apoptosis and cell-cycle regulation. Colon biopsies showed no histological abnormalities by haematoxylin and eosin staining. Immunohistochemical staining, however, detected NF-kappaB activation in tissue macrophages using antibodies to the nuclear localization sequence for p65 and the F4/80 marker for murine macrophages. Similarly, haematin-starved E. faecalis strongly activated NF-kappaB in murine macrophages in vitro. Furthermore, primary and transformed colonic epithelial cells activated the G2/M checkpoint in vitro following exposure to haematin-starved E. faecalis. Modulation of this cell-cycle checkpoint was due to extracellular superoxide produced as a result of the respiratory block in haematin-starved E. faecalis. These results demonstrate that the uniquely dichotomous metabolism of E. faecalis can significantly modulate gene expression in the colonic mucosa for pathways associated with inflammation, apoptosis and cell-cycle regulation.
Journal of Medical Microbiology 11/2008; 57(Pt 10):1193-204. · 2.50 Impact Factor
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ABSTRACT: Blood borne Listeria monocytogenes enter the CNS via migration of parasitized Ly-6Chigh monocytes, but the signals that trigger this migration are not known. To understand more completely events leading to monocyte recruitment, experiments presented here combined microarray analysis of gene expression in the brains of experimentally infected mice with measurements of bacterial CFU and serum cytokines following i.v. infection with L. monocytogenes. At 24 and 48 h postinfection, the brain was sterile but there were significant changes in transcriptional activity related to serum proinflammatory cytokines. Real-time PCR confirmed mRNA up-regulation of genes related to IFN-gamma, IL-1, and TNF-alpha, although IFN-gamma itself was not up-regulated in the brain. Infection with Deltaacta, but not Deltahly mutants, increased serum concentrations of IFN-gamma, IL-6, and to a lesser extent TNF-alpha. The brain was not infected but there was widespread mRNA up-regulation in it and an influx of Ly-6Chigh monocytes in Deltaacta-infected mice. Moreover, DeltaactA-infected IFN-gamma-/- mice had no brain influx of Ly-6Chigh monocytes despite normal monocyte trafficking from bone marrow to blood and spleen. Additionally, IFN-gamma-/- mice showed diminished mRNA expression for monocyte-attracting chemokines, and significantly less CXCL9 and CXCL10 protein in the brain compared with normal mice. These data demonstrate that monocyte recruitment to the brain is independent of bacterial invasion of the CNS and is triggered by proinflammatory cytokines, in particular IFN-gamma, produced by the innate immune response to intracellular infection in peripheral organs.
The Journal of Immunology 08/2008; 181(1):529-36. · 5.79 Impact Factor
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Jay S Hanas,
James R Hocker,
John Y Cheung,
Jason L Larabee, Megan R Lerner,
Stan A Lightfoot,
Daniel L Morgan,
Kent D Denson,
Kristi C Prejeant,
Yuiry Gusev,
Brenda J Smith,
Rushie J Hanas,
Russell G Postier,
Daniel J Brackett
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ABSTRACT: The aim of this study is to identify biomarkers in sera of pancreatic cancer patients using mass spectrometry (MS) approaches.
Sera from patients diagnosed with pancreatic adenocarcinoma and sera from normal volunteers were subjected to gel electrophoresis to resolve and quantify differences in protein levels. Protein bands that differed quantitatively were digested with trypsin, and peptides were identified by electrospray ionization (ESI) ion-trap tandem MS. Mass spectra were also collected directly from pancreatic cancer sera as well as healthy control sera using ESI-MS.
Three large-mass proteins were found to be elevated in pancreatic cancer sera versus normal sera, alpha-2 macroglobulin, ceruloplasmin, and complement 3C. Complement 3C is a major regulator of inflammatory responses. The ESI-MS of human pancreatic cancer sera versus normal sera revealed greater heterogeneity in cancer sera than control sera, especially in the low-mass region. Bootstrapping statistical analysis identified 20 low-mass serum peaks that correlated with control sera and 20 different peaks that correlated with pancreatic cancer sera.
The fact that inflammation-sensitive proteins were identified as increased in pancreatic cancer sera supports the hypothesis that inflammatory-driven processes are involved in pancreatic carcinogenesis. Liquid ESI-MS analyses of sera hold promise for future pancreatic cancer blood tests as well as for understanding mechanisms of pancreatic carcinogenesis. The variability observed between the low-mass regions of normal versus pancreatic cancer spectra may aid in diagnosis and therapy.
Pancreas 02/2008; 36(1):61-9. · 2.39 Impact Factor
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ABSTRACT: Visually guided ocular growth is facilitated by scleral extracellular matrix remodeling at the posterior pole of the eye. Coincident with scleral remodeling, significant changes in choroidal morphology, blood flow, and protein synthesis have been shown to occur in eyes undergoing ocular growth changes. The current study is designed to identify gene expression changes that may occur in the choroid/retinal pigment epithelium (RPE) of marmoset eyes during their compensation for hyperopic defocus as compared to eyes compensating for myopic defocus.
Total RNA was isolated from choroid/RPE from four common marmosets (Callithrix jacchus) undergoing binocular lens treatment using extended wear soft contact lenses of equal magnitude but opposite sign (+/-5 diopter [D]). After reverse transcription, cDNA was labeled and hybridized to a human oligonucleotide microarray and gene transcript expression profiles were determined. Real-time polymerase chain reaction (PCR) and western blot analysis were used to confirm genes and proteins of interest, respectively.
Microarray analyses in choroid/RPE indicated 204 genes were significantly changed in minus lens-treated as compared with plus lens-treated eyes (p<0.05, Student's t-test). Differential choroid/RPE expression of protein tyrosine phosphatase, receptor type, B (PTPRB), transforming growth factor beta-induced (TGFBI), and basic fibroblast growth factor 2 (FGF-2) were confirmed by real-time PCR. TGFBIp was confirmed at the protein level by western blot analysis in marmoset and human cornea, choroid/RPE, and sclera.
The present study demonstrated that significant gene expression changes occur in the marmoset choroid/RPE during visually guided ocular growth. The identification of novel candidate genes in choroid/RPE of marmoset eyes actively accelerating or decelerating their rates of ocular elongation may elucidate the choroidal response during the regulation of postnatal ocular growth and may lead to the identification of choroid/RPE signaling molecules that participate in scleral remodeling.
Molecular vision 01/2008; 14:1465-79. · 2.20 Impact Factor
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ABSTRACT: Epidemiological and clinical evidence indicate that inflammatory processes play a pivotal role in a number of conditions associated with aging, including osteoporosis and cardiovascular diseases. The purpose of this study was to evaluate cardiovascular pathology and select inflammatory mediators of interest in a model of low-grade inflammation-induced osteopenia.
Slow-release pellets were subcutaneously implanted in male rats to deliver 0, 3.3, or 33.3 microg of lipopolysaccharide (LPS)/day for 90 days. Tail blood was collected at 1, 2, and 3 months for differential white cell counts, and at the end of the study, hearts were harvested for histological and immunohistochemical evaluation.
The low-grade inflammatory response was characterized by elevated peripheral blood neutrophils and monocytes. Histological examination of heart cross sections revealed increased fibrous tissue, infiltration of lymphocytes, accumulation of mast cells, and roughened intimal borders within the arteries and arterioles, consistent with vascular disease. Inflammatory mediators (cyclooxygenase-2, tumor necrosis factor-alpha, and interleukin-1 beta) were up-regulated, and increased expression of platelet endothelial cell adhesion molecule-1 and receptor activator for NF-kappaB ligand was localized to the microvasculature endothelium.
These findings suggest that inflammation induced by chronic exposure to LPS produces cardiovascular pathology in the smaller intramural arteries and arterioles and support the utility of this model for further mechanistic and therapeutic studies focused on the role of chronic inflammation in cardiovascular disease.
Cardiovascular pathology: the official journal of the Society for Cardiovascular Pathology 11/2007; 18(1):1-10. · 1.63 Impact Factor
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ABSTRACT: microRNAs are functional, 22 nt, noncoding RNAs that negatively regulate gene expression. Disturbance of microRNA expression may play a role in the initiation and progression of certain diseases. A microRNA expression signature has been identified that is associated with pancreatic cancer. This has been accomplished with the application of real-time PCR profiling of over 200 microRNA precursors on specimens of human pancreatic adenocarcinoma, paired benign tissue, normal pancreas, chronic pancreatitis and nine pancreatic cancer cell lines. Hierarchical clustering was able to distinguish tumor from normal pancreas, pancreatitis and cell lines. The PAM algorithm correctly classified 28 of 28 tumors, 6 of 6 normal pancreas and 11 of 15 adjacent benign tissues. One hundred microRNA precursors were aberrantly expressed in pancreatic cancer or desmoplasia (p < 0.01), including microRNAs previously reported as differentially expressed in other human cancers (miR-155, miR-21, miR-221 and miR-222) as well as those not previously reported in cancer (miR-376a and miR-301). Most of the top aberrantly expressed miRNAs displayed increased expression in the tumor. Expression of the active, mature microRNA was validated using a real-time PCR assay to quantify the mature microRNA and Northern blotting. Reverse transcription in situ PCR showed that three of the top differentially expressed miRNAs (miR-221, -376a and -301) were localized to tumor cells and not to stroma or normal acini or ducts. Aberrant microRNA expression may offer new clues to pancreatic tumorigenesis and may provide diagnostic biomarkers for pancreatic adenocarcinoma.
International Journal of Cancer 03/2007; 120(5):1046-54. · 5.44 Impact Factor
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ABSTRACT: Evidence from epidemiological, clinical and animal studies suggests a link may exist between low bone density and cardiovascular disease, with inflammatory mediators implicated in the pathophysiology of both conditions. This project examined whether supplementation with soy isoflavones (IF), shown to have anti-inflammatory properties, could prevent tissue expression of TNF-alpha and the development of skeletal pathology in an animal model of chronic inflammation.
Eight-week old, intact, female C57BL/6J mice were used. In Phase 1, a lipopolysaccharide (LPS)-dose response study (0, 0.133, 1.33 and 13.3 mug/d) was conducted to determine the LPS dose to use in Phase 2. The results indicated the 1.33 mug LPS/d dose produced the greatest decrease in lymphocytes and increase in neutrophils. Subsequently, in Phase 2, mice were randomly assigned to one of six groups (n = 12-13 per group): 0 or 1.33 mug LPS/d (placebo or LPS) in combination with 0, 126 or 504 mg aglycone equivalents of soy IF/kg diet (Control, Low or High dose IF). Mice were fed IF beginning 2 wks prior to the 30-d LPS study period.
At the end of the study, no differences were detected in final body weights or uterine weights. In terms of trabecular bone microarchitecture, muCT analyses of the distal femur metaphysis indicated that LPS significantly decreased trabecular bone volume (BV/TV) and number (TbN), and increased separation (TbSp). Trabecular bone strength (i.e. total force) and stiffness were also compromised in response to LPS. The High IF dose provided protection against these detrimental effects on microarchitecture, but not biomechanical properties. No alterations in trabecular thickness (TbTh), or cortical bone parameters were observed in response to the LPS or IF. Immunohistomchemical staining showed that tumor necrosis factor (TNF)-alpha was up-regulated by LPS in the endothelium of small myocardial arteries and arterioles as well as the tibial metaphysis and down-regulated by IF.
These results suggest IF may attenuate the negative effects of chronic inflammation on bone and cardiovascular health. Additional research is warranted to examine the anti-inflammatory properties of the soy isoflavones and the mechanisms underlying their prevention of chronic inflammation-induced bone loss.
Journal of Inflammation 02/2007; 4:17. · 2.26 Impact Factor
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ABSTRACT: In vitro stable isotope glucose tracer studies indicate that undifferentiated cells of the pancreas use glucose primarily through the nonoxidative reactions of the pentose cycle for nucleic acid ribose synthesis, whereas normal or less transformed cells primarily use the oxidative branch of the cycle.
The pancreatic heads of 4 groups (5/group) of male rats were implanted with time-release pellets designed to deliver placebo or 7,12-dimethylbenzanthracene (DMBA) at 11, 33, or 56 mg/d. Four weeks after pancreatic exposure to DMBA, [1,2-C2]-D-glucose tracer (1 g/kg) was injected intraperitoneally followed by sera collection at 1 and 2 hours and harvest of tumors, adjacent pancreatic tissue, and sera at 3 hours.
Tumors (2-9 mm) were found across DMBA groups, with the largest in the high-dose group (> or =5 mm). Selective monitoring by gas chromatography-mass spectrometry of the doubly-labeled [1,2-C2]-D-ribose of RNA, which requires nonoxidative synthesis in the pentose cycle, showed a 2.8-, 2.9-, and 5.7-fold increase in pancreatic tumors. Liver and adjacent pancreas preferentially produced [1-C1]-D-ribose through the oxidative reactions of the cycle. Tumor-bearing animals also cleared and recycled tracer glucose at a faster rate.
Simultaneous selective positional ion monitoring of C-labeled metabolites and their mass isotopomers in tissues and blood opens new avenues for the early detection and response to therapy testing of pancreatic cancer using GC-MS and/or magnetic resonance imaging-based methods. This study emphasizes the benefits of stable isotope-based dynamic metabolic profiling, when applied in vivo, and the several advantages it offers to positron emission tomography.
Pancreas 12/2005; 31(4):337-43. · 2.39 Impact Factor
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ABSTRACT: The purpose of this study was to establish experimental conditions to produce apoptosis by the fluorinated pyrimidine 5-fluorouridine and to examine the changes in gene expression that occurred during cell death. HCT-116 colorectal carcinoma cells were exposed to 10 microM 5-fluorouridine alone or in the presence of 1 mM uridine, 30 microM thymidine or both uridine and thymidine. A time-dependent increase in the percentage of apoptotic cells and a decrease in the percentage of viable cells were observed when the cells were treated with 5-fluorouridine in the absence of uridine (p < 0.001) but not in the presence of uridine. cDNA microarray analysis was used to study the expression of 1,200 different genes during apoptosis by 5-flurouridine. The expression of 33 genes was upregulated by 5-fold or greater at 16 and 24 h of 5-fluorouridine exposure. The largest cluster of upregulated genes included a group of genes classified as growth factors, cytokines and chemokines (e.g. interleukin-3, interleukin-4, B-cell growth factor 1 and stem cell growth factor). The expression of MIC-1 increased up to 100-fold during 5-flurouridine exposure. One hundred and twenty-four genes were downregulated by 5-fold or greater following exposure to 5-fluorouridine. The downregulated genes were distributed throughout the six different classifications on the array. Our data demonstrate a diverse pattern of gene expression during the fluorouridine-induced apoptosis and suggest that mechanisms besides a global inhibition of RNA synthesis/ processing contribute to the RNA-directed cytotoxicity of fluoropyrimidines.
International Journal of Oncology 09/2005; 27(2):297-306. · 2.40 Impact Factor
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ABSTRACT: Sigma receptors have recently been implicated in the actions of cocaine, and antagonists of these receptors prevent many acute and subchronic cocaine effects. A previous study revealed that the immediate early gene fra-2 is up-regulated after cocaine administration, and this effect is prevented by the sigma-1 receptor antagonist BD1063 [1-[2-(3,4-dichlorophenyl)ethyl]-4-methylpiperazine]. In the present study, the effects of cocaine and BD1063 on the expression of six fos and jun genes were evaluated in mouse brains using cDNA microarrays. Several of these genes were altered by cocaine, but only the alteration in fra-2 was prevented by BD1063. The time courses of fra-2 and sigma-1 receptor gene and protein expression in different brain regions were also determined. Cocaine up-regulated fra-2, which was followed by a later up-regulation of sigma-1 receptors. The cocaine-induced up-regulation of fra-2 and sigma-1 receptor genes and proteins were detected in whole brain, striatum, and cortex, but not in cerebellum. All of these cocaine-induced effects were prevented by BD1063. The interaction between cocaine, fra-2, and sigma-1 receptors involves brain regions that are established components of the neural circuit for reward, suggesting that they may contribute to the enduring changes that underlie the cellular basis of drug abuse.
Journal of Pharmacology and Experimental Therapeutics 09/2005; 314(2):770-9. · 3.83 Impact Factor
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ABSTRACT: The prohibitin 3' untranslated region (3'UTR) belongs to a novel class of non-coding regulatory RNAs. It arrests cell cycle progression by blocking G1-S transition in breast and other cancers. Our previous studies comparing MCF7 derived clones constitutively expressing a common allelic form of prohibitin RNA (UTR/C) to various controls demonstrated that it functions as a tumor suppressor. Here, we further characterized the morphology and motility of these transgenic breast cancer cells when grown in cell culture and on nude mice. In contrast to empty vector (EV) cells, UTR/C cells were observed to grow in an organized manner with more cell-cell contact and differentiate into structures with a duct-like appearance. Computer assisted cytometry to evaluate differences in nuclear morphology was performed on UTR/C and EV tissues from nude mice. Receiver operator curve areas generated using a logistic regression model were 0.8, indicating the ability to quantitatively distinguish UTR/C from EV tissues. Keratinocyte growth factor-induced motility experiments showed that migration of UTR/C cells was significantly reduced (80-90%) compared to EV cells. Together, these data indicate that this novel 3'UTR influences not only the tumorigenic phenotype but also may play a role in differentiation and migration of breast cancer cells.
Journal of Molecular Histology 09/2004; 35(6):639-46. · 1.48 Impact Factor
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ABSTRACT: A significant component of the general surgeon's practice is treatment of cancer that most commonly involves breast, colorectal, pancreas, and hepatic surgeries. The prognosis for these patients is largely dependent upon the stage of the disease at the time of diagnosis. There is convincing evidence that the induction and progression of cancer occurs when the function of genes that regulate cell proliferation, cell death and DNA repair becomes altered. These molecular disturbances result from ongoing, tumor-promoting activity long before symptoms of the disease become apparent. The role of many genes and the extent of their involvement in this deleterious, molecular process is unique to the organ. However, the tumor suppressor genes p53 and p16, the oncogene c-myc, and the inflammation-associated enzyme COX-2 are implicated in all four of the tumors. The roles of the oncogenes ras and HER-2/erbB-2/neu are significant in all of the tumors except hepatocellular carcinoma. The systematic establishment of a comprehensive data base relating gene expression activity to the initiation and development of cancer will provide molecular markers for detection and targets for treatment. This overview presents the current status of information associating gene expression activity with cancers typically encountered by general surgeons.
The Journal of the Oklahoma State Medical Association 11/2003; 96(10):485-94.
