Transcriptional changes in U343 MG-a glioblastoma cell line exposed to ionizing radiation

Glioblastoma multiforme (GBM) is a highly invasive and radioresistant brain tumor. Aiming to study how glioma cells respond to gamma-rays in terms of biological processes involved in cellular responses, we performed experiments at cellular context and gene expression analysis in U343-MG-a GBM cells irradiated with 1 Gy and collected at 6 h post-irradiation. The survival rate was approximately 61% for 1 Gy and was completely reduced at 16 Gy. By performing the microarray technique, 859 cDNA clones were analyzed. The Significance Analysis of Microarray algorithm indicated 196 significant expressed genes (false discovery rate (FDR) = 0.42%): 67 down-regulated and 97 up-regulated genes, which belong to several classes: metabolism, adhesion/cytoskeleton, signal transduction, cell cycle/apoptosis, membrane transport, DNA repair/DNA damage signaling, transcription factor, intracellular signaling, and RNA processing. Differential expression patterns of five selected genes (HSPA9B, INPP5A, PIP5K1A, FANCG, and TPP2) observed by the microarray analysis were further confirmed by the quantitative real time RT-PCR method, which demonstrated an up-regulation status of those genes. These results indicate a broad spectrum of biological processes (which may reflect the radio-resistance of U343 cells) that were altered in irradiated glioma cells, so as to guarantee cell survival.

Stable and high-level production of recombinant Factor IX in human hepatic cell line

Hemophilia B is a genetic disease of the coagulation system that affects one in 30,000 males worldwide. Recombinant human Factor IX (rhFIX) has been used for hemophilia B treatment, but the amount of active protein generated by these systems is inefficient, resulting in a high-cost production of rhFIX. In this study, we developed an alternative for rhFIX production. We used a retrovirus system to obtain two recombinant cell lines. We first tested rhFIX production in the human embryonic kidney 293 cells (293). Next, we tested a hepatic cell line (HepG2) because FIX is primarily expressed in the liver. Our results reveal that intracellular rhFIX expression was more efficient in HepG2/rhFIX (46%) than in 293/rhFIX (21%). The activated partial thromboplastin time test showed that HepG2/rhFIX expressed biologically active rhFIX 1.5 times higher than 293/rhFIX (P = 0.016). Recovery of rhFIX from the HepG2 by reversed-phase chromatography was straightforward. We found that rhFIX has a pharmacokinetic profile similar to that of FIX purified from human plasma when tested in hemophilic B model. HepG2/rhFIX cell line produced the highest levels of rhFIX, representing an efficient in vitro expression system. This work opens up the possibility of significantly reducing the costs of rhFIX production, with implications for expanding hemophilia B treatment in developing countries.

Pediatric glioblastoma cell line shows different patterns of expression of transmembrane ABC transporters after in vitro exposure to vinblastine

Resistance to drug is a major cause of treatment failure in pediatric brain cancer. The multidrug resistance (MDR) phenotype can be mediated by the superfamily of adenosine triphosphate-binding cassette (ABC) transporters. The dynamics of expression of the MDR genes after exposure to chemotherapy, especially the comparison between pediatric brain tumors of different histology, is poorly described. To compare the expression profiles of the multidrug resistance genes ABCB1, ABCC1, and ABCG2 in different neuroepithelial pediatric brain tumor cell lines prior and following short-term culture with vinblastine. Immortalized lineages from pilocytic astrocytoma (R286), anaplasic astrocytoma (UW467), glioblastoma (SF188), and medulloblastoma (UW3) were exposed to vinblastine sulphate at different schedules (10 and 60 nM for 24 and 72 h). Relative amounts of mRNA expression were analyzed by real-time quantitative polymerase chain reaction. Protein expression was assessed by immunohistochemistry for ABCB1, ABCC1, and ABCG2. mRNA expression of ABCB1 increased together with augmenting concentration and time of exposure to vinblastine for R286, UW467, and UW3 cell lines. Interestingly, ABCB1 levels of expression diminished in SF188. Following chemotherapy, mRNA expression of ABCC1 decreased in all cell lines other than glioblastoma. ABCG2 expression was influenced by vinblastine only for UW3. The mRNA levels showed consistent association to protein expression in the selected sets of cell lines analyzed. The pediatric glioblastoma cell line SF188 shows different pattern of expression of multidrug resistance genes when exposed to vinblastine. These preliminary findings may be useful in determining novel strategies of treatment for neuroepithelial pediatric brain tumors.

Cytotoxic effects of butanolic extract from Pfaffia paniculata (Brazilian Ginseng) on cultured human breast cancer cell line MCF-7

Roots of Pfaffia paniculata have been well documented for multifarious therapeutic values and have also been used for cancer therapy in folk medicine. This study has been performed in a human breast tumor cell line, the MCF-7 cells. These are the most commonly used model of estrogen-positive breast cancer, and it has been originally established in 1973 at the Michigan Cancer Foundation from a pleural effusion taken from a woman with metastatic breast cancer. Butanolic extract of the roots of P. paniculata showed cytotoxic effect MCF-7 cell line. as determined with crystal violet assay, cellular death with acridine orange/ethidium bromide staining, and cell proliferation with immunocytochemistry of bromodeoxyuridine (BrdU). Subcellular alterations were evaluated by electron microscopy. Cells treated With butanolic extract showed degeneration of cytoplasmic components and profound morphological and nuclear alterations. The results show that this butanolic extract indeed presents cytotoxic substances, and its fractions merit further investigations. (C) 2008 Elsevier GmbH. All rights reserved.

Acid-Base Changes in Canine Neonates Following Normal Birth or Dystocia

There are limited data concerning blood gas parameters in neonatal dogs. Knowledge of the normal physiology may facilitate effective therapeutic intervention and potentially reduce neonatal mortality. This study examined acid-base parameters in pups born at normal parturition (n = 27) compared with those born after obstetrical assistance or caesarean operation (n = 13) and those born following oxytocin (OXY) administration for treatment of uterine inertia (n = 11). Pups were subjected to an objective scoring method of neonatal health adapted from use in humans (the Apgar score) at birth and again at 5 and 60 min after birth. Venous blood samples were collected at 5 and 60 min after birth for evaluation of blood gas parameters. At birth, all pups had low Apgar scores and a mixed acidosis. The base excess was lowest for pups delivered after OXY administration. The Apgar score improved for all pups after 5 min of birth and there was an improvement in carbon dioxide tension, base excess and venous blood pH at 1 h, although in all pups a metabolic acidosis persisted. These data provide an important insight into neonatal physiology and the variability of blood gas parameters in pups born at normal and abnormal parturition and provide the basis for clinical decision making following dystocia.

Effects of Ethanol on the Surface and Bulk Properties of a Microwave-Processed PMMA Denture Base Resin

Purpose: This study evaluated the effect of different concentrations of ethanol on hardness, roughness, flexural strength, and color stability of a denture base material using a microwave-processed acrylic resin as a model system. Materials and Methods: Sixty circular (14 x 4 mm) and 60 rectangular microwave-polymerized acrylic resin specimens (65 x 10 x 3 mm(3)) were employed in this study. The sample was divided into six groups according to the ethanol concentrations used in the immersion solution, as follows: 0% (water), 4.5%, 10%, 19%, 42%, and 100%. The specimens remained immersed for 30 days at 37 degrees C. The hardness test was performed by a hardness tester equipped with a Vickers diamond penetrator, and a surface roughness tester was used to measure the surface roughness of the specimens. Flexural strength testing was carried out on a universal testing machine. Color alterations (Delta E) were measured by a portable spectrophotometer after 12 and 30 days. Variables were analyzed by ANOVA/Tukey`s test (alpha = 0.05). Results: For the range of ethanol-water solutions for immersion (water only, 4.5%, 10%, 19.5%, 42%, and 100%), the following results were obtained for hardness (13.9 +/- 2.0, 12.1 +/- 0.7, 12.9 +/- 0.9, 11.2 +/- 1.5, 5.7 +/- 0.3, 2.7 +/- 0.5 VHN), roughness (0.13 +/- 0.01, 0.15 +/- 0.07, 0.13 +/- 0.05, 0.13 +/- 0.02, 0.23 +/- 0.05, 0.41 +/- 0.19 mu m), flexural strength (90 +/- 12, 103 +/- 18, 107 +/- 16, 90 +/- 25, 86 +/- 22, 8 +/- 2 MPa), and color (0.8 +/- 0.6, 0.8 +/- 0.3, 0.7 +/- 0.4, 0.9 +/- 0.3, 1.3 +/- 0.3, 3.9 +/- 1.5 Delta E) after 30 days. Conclusions: The findings of this study showed that the ethanol concentrations of tested drinks affect the physical properties of the investigated acrylic resin. An obvious plasticizing effect was found, which could lead to a lower in vivo durability associated with alcohol consumption.

The Effect of Polymerization Cycles on Color Stability of Microwave-Processed Denture Base Resin

Purpose: This study evaluated the effect of different microwave polymerization cycles on the color changes of a microwave-processed denture base resin after accelerated aging and immersion in beverages. Materials and Methods: Specimens of light pink acrylic resin were divided into three groups according to polymerization cycle: (A) 500 W for 3 minutes, (B) 90 W for 13 minutes + 500 W for 90 seconds, and (C) 320 W for 3 minutes + 0 W for 4 minutes + 720 W for 3 minutes. Control groups were a heat-processed acrylic resin (T) and a chemically activated denture repair resin (Q). Eight specimens per group were aged in an artificial aging chamber and evaluated at 20, 192, and 384 hours. Another series of 40 specimens per group were immersed in water, coffee, tea, cola, or red wine and evaluated at 1, 12, and 36 days. Color was measured by a spectrophotometer before and after aging or immersion. Color changes (Delta E) were analyzed by ANOVA/Bonferroni t-test (alpha = 0.05). Results: Mean Delta E (+/- SD) after 384 hours of accelerated aging were (A) 2.51 +/- 0.50; (B) 3.16 +/- 1.09; (C) 2.89 +/- 1.06; (T) 2.64 +/- 0.34; and (Q) 9.03 +/- 0.40. Group Q had a significantly higher Delta E than the other groups. Color changes of immersed specimens were significantly influenced by solutions and time, but the five groups showed similar values. Mean Delta E at 36 days were (water) 1.4 +/- 0.8; (coffee) 1.3 +/- 0.6; (tea) 1.7 +/- 0.5; (cola) 1.4 +/- 0.7; and (red wine) 10.2 +/- 2.7. Results were similar among the five test groups. Conclusions: Color changes of the microwave-polymerized denture base resin tested were not affected by different polymerization cycles after accelerated aging or immersion in beverages. These changes were similar to the conventional heat-polymerized acrylic resin test, but lower than the repair resin after accelerated aging.

Evaluation of the Bond Strength of Denture Base Resins to Acrylic Resin Teeth: Effect of Thermocycling

Purpose: The purpose of this study was to evaluate the thermocycling effects and shear bond strength of acrylic resin teeth to denture base resins. Materials and Methods: Three acrylic teeth (Biotone, Trilux, Ivoclar) were chosen for bonding to four denture base resins: microwave-polymerized (Acron MC), heat-polymerized (Lucitone 550 and QC-20), and light-polymerized (Versyo. bond). Twenty specimens were produced for each denture base/acrylic tooth combination and were divided into two groups (n = 10): without thermocycling (control groups) and thermocycled groups submitted to 5000 cycles between 4 and 60 degrees C. Shear strength tests (MPa) were performed with a universal testing machine at a crosshead speed of 1 mm/min. Statistical analysis of the results was carried out with three-way ANOVA and Bonferroni`s multiple comparisons post hoc analysis for test groups (alpha = 0.05). Results: The shear bond strengths of Lucitone/Biotone, Lucitone/Trilux, and Versyo/Ivoclar specimens were significantly decreased by thermocycling, compared with the corresponding control groups (p < 0.05). The means of Acron/Ivoclar and Lucitone/Ivoclar specimens increased after thermocycling (p < 0.05). The highest mean shear bond strength value was observed with Lucitone/Biotone in the control group (14.54 MPa) and the lowest with QC-20/Trilux in the thermocycled group (3.69 MPa). Conclusion: Some acrylic tooth/denture base resin combinations can be more affected by thermocycling; effects vary based upon the materials used.

Effect of methyl methacrylate monomer on bond strength of denture base resin to acrylic teeth

Bond failures at the acrylic teeth and denture base resin interface are still a common clinical problem in prosthodontics. The effect of methyl methacrylate (MMA) monomer on the bond strength of three types of denture base resins (Acron MC, Lucitone 550 and QC-20) to two types of acrylic teeth (Biotone and Trilux) was evaluated. Twenty specimens were produced for each denture base resin/acrylic tooth combination and were randomly divided into control (acrylic teeth received no surface treatment) and experimental groups (MMA was applied to the surface of the acrylic teeth for 180 s) and were submitted to shear tests (1 mm/mm). Data (MPa) were analyzed using three-way ANOVA/Student`s test (alpha = 0.05). MMA increased the bond strength of Lucitone denture base resins and decreased the bond strength of QC-20. No difference was detected for the bond strength of Acron MC base resin after treatment with MMA. (C) 2008 Elsevier Ltd. All rights reserved.