cDNA microarray reveals altered cytoskeletal gene expression in space-flown leukemic T lymphocytes (Jurkat).
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Cell Division↗
Biomedical subjects
E H Piepmeier
Publications and source records attributed to E H Piepmeier.
Exposure to hyperbaric oxygen induces cell cycle perturbation in prostate cancer cells.
Cell cycle synchronization of tumor cells by exposure to hyperbaric oxygenation (HBO) may increase the efficacy of chemotherapy or radiation by placing cells into a chemosensitive portion of the cycle. The purpose of the current study was to examine oxygen pressure-dependent relationships with respect to the cell cycle in prostate tumor cells in vitro. LNCaP cells were grown in an incubator at 21% O2 and then exposed to 100% oxygen at pressures up to 6 atmospheres (atm) for 1.5 h. Cells were then returned to the incubator and evaluated for DNA content by propidium iodide and new DNA synthesis with a pulse-chase experiment. Cell cycle effects were evaluated by flow cytometry. Exposure to HBO increased the percentage of cells synthesizing new DNA in a dose-dependent fashion: 0 atm, 44%; 6 atm, 65%. Cells that synthesize new DNA accumulate in G2/M as a function of partial pressure of oxygen. These results suggest that HBO induces cells to enter the cell cycle and accumulate in G2/M. Cell cycle synchronization and entry of senescent cells into the cell cycle suggest that HBO may be a useful adjuvant to chemotherapy or radiation in the treatment of prostate cancer. There are two potential mechanisms of action that may make HBO efficacious in the treatment of prostate cancer. HBO may potentiate cancer chemotherapeutic agents that cause damage to DNA during DNA synthesis or HBO may inhibit cell division causing accumulation in G2/M.
Atmospheric Pressure↗
Fibroblast response to rapid decompression and hyperbaric oxygenation.
INTRODUCTION: The cellular basis for symptoms associated with rapid decompression and the use of hyperbaric oxygenation treatment (HBO) is not established. METHODS: Image analysis, sulforhodamine B assay and bromodeoxyuridine (BrDU) incorporation were used to identify cellular changes associated with rapid decompression (RD) or hyperbaric oxygenation (HBO). Human fibroblasts were exposed to RD or HBO and compared with untreated cells. Immediately following treatment, the fraction of cells synthesizing DNA was measured by detection of cells incorporating the BrDU. At 1 d and 3 d following the treatments, total cell protein adherent to the bottom of wells was measured using a sulforhodamine B assay. Cell density was observed with light microscopy and quantified with image analysis. RESULTS: RD increased total protein significantly, (p < 0.05) relative to control, while HBO had less effect. The fraction of cells synthesizing DNA was increased by HBO and reduced by RD relative to control (p < 0.05). Image analysis showed that cell density at day 1 was: control>HBO>RD; and at day 3: RD>HBO>control, indicating an increase in proliferation induced by the treatments. CONCLUSION: This data shows that HBO and RD increase the proliferation of fibroblasts for 24 h following treatment. HBO increased DNA replication. While there was a decrease in DNA replication following RD, protein synthesis was enhanced.
Bromodeoxyuridine↗
Spaceflight alters microtubules and increases apoptosis in human lymphocytes (Jurkat).
Alteration in cytoskeletal organization appears to underlie mechanisms of gravity sensitivity in space-flown cells. Human T lymphoblastoid cells (Jurkat) were flown on the Space Shuttle to test the hypothesis that growth responsiveness is associated with microtubule anomalies and mediated by apoptosis. Cell growth was stimulated in microgravity by increasing serum concentration. After 4 and 48 h, cells filtered from medium were fixed with formalin. Post-flight, confocal microscopy revealed diffuse, shortened microtubules extending from poorly defined microtubule organizing centers (MTOCs). In comparable ground controls, discrete microtubule filaments radiated from organized MTOCs and branched toward the cell membrane. At 4 h, 30% of flown, compared to 17% of ground, cells showed DNA condensation characteristic of apoptosis. Time-dependent increase of the apoptosis-associated Fas/ APO-1 protein in static flown, but not the in-flight 1 g centrifuged or ground controls, confirmed microgravity-associated apoptosis. By 48 h, ground cultures had increased by 40%. Flown populations did not increase, though some cells were cycling and actively metabolizing glucose. We conclude that cytoskeletal alteration, growth retardation, and metabolic changes in space-flown lymphocytes are concomitant with increased apoptosis and time-dependent elevation of Fas/APO-1 protein. We suggest that reduced growth response in lymphocytes during spaceflight is linked to apoptosis.
Apoptosis↗
Prolonged weightlessness affects promyelocytic multidrug resistance.
An immortalized promyelocytic cell line was studied to detect how doxorubicin uptake is affected by microgravity. The purpose of this experiment was to identify the effect that microgravity may have on multidrug resistance in leukocytes. HL60 cells and HL60 cells resistant to anthracycline (HL60/AR) were grown in RPMI and 10% FBS. Upon reaching orbit in the Space Shuttle Endeavour, the cells were robotically mixed with doxorubicin. Three days after mixing, cells were fixed with paraformaldehyde/glutaraldehyde. Ground control experiments were conducted concurrently using a robot identical to the one used on the Shuttle. Fixed cells were analyzed within 2 weeks of launch. Confocal micrographs identified changes in cell structure (transmittance), drug distribution (fluorescence), and microtubule polymerization (fluorescence). Flight cells showed a lack of cytoskeletal polymerization resulting in an overall amorphic globular shape. Doxorubicin distribution in ground cells included a large numbers of vesicles relative to flight cells. There was a greater amount of doxorubicin present in flight cells (85% +/- 9.7) than in ground control cells (43% +/- 26) as determined by image analysis. Differences in microtubule formation between flight cells and ground cells could be partially responsible for the differences in drug distribution. Cytoskeletal interactions are critical to the function of P-glycoprotein as a drug efflux pump responsible for multidrug resistance.
Biological Transport↗
Effect of dextran 70 blood substitute on diazepam binding by human serum albumin.
Diazepam binding studies with dextran 70 and human serum albumin (HSA) were carried out using centrifugation and a membrane ultrafiltration technique to separate the drug protein complex and the free microsolute. The molecular filtration method was found to be precise and reproducible with very little variations. The binding of diazepam to dextran was significant but considerably smaller when compared with the extensive binding of diazepam to HSA. The results of these studies demonstrate that diazepam binding of HSA is altered in the presence of dextran. Diazepam binds to dextran resulting in displacement of diazepam from albumin binding sites. The contributing effect of dextran to overall diazepam binding increased as the dextran concentration increased and the HSA concentration decreased. However, the net result of HSA dilution with dextran 70 was still an increase in the percent free diazepam. Various concentrations of dextran 70 significantly displaced HSA bound diazepam in all the mixtures studied.
Journal Article↗
The influence of hyperbaric oxygenation on leukocyte viability and surface protein expression.
Hyperbaric oxygenation was studied as a potential inducer of cell growth and differentiation in promyelocytic leukemic HL60 cells. We studied changes in HL60 cells exposed to hyperbaric oxygen, oxygen, or carbon dioxide for 72 h. The proliferation rate and viability of cells in the hyperbaric oxygenation groups were significantly lower (p < 0.05) than for the controls. While CD13 and CD38 were expressed less following hyperbaric treatment, CD11b, CD14, and CD16 showed an increase following hyperbaric treatment, and CD10, CD15, and HLADR showed no change. These results support previous studies which demonstrate the role of oxygen tension in the regulation of cell cycle and protein expression.
Antigens, CD↗
Liquid-interfaced oscillating glow discharge detector for a flowing liquid system.
A new liquid-interfaced oscillating glow discharge detector having a frequency and current response to femtomole and picomole quantities respectively of potassium nitrate and sucrose injected into an aqueous flowing eluent is presented. The glow discharge is formed in an argon atmosphere at ambient pressure between a platinum anode and a cathode consisting of an aqueous conducting solution. A detailed description of the appearance of the liquid-interfaced glow discharge at various electrode distances and the occurrence of high-frequency oscillations is given.
Argon↗
Atmospheric pressure effects on glucose monitoring devices.
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Atmospheric Pressure↗
Evaluation of glucose monitoring devices in the hyperbaric chamber.
Diabetic patients may experience fluctuations in whole blood glucose (WBG) levels while receiving hyperbaric oxygen therapy (HBO) resulting in seizure-like activity. Therefore, hyperbaric medical attendants must accurately monitor the WBG levels of these patients during HBO. In addressing this concern, this study evaluated the accuracy and reliability of commercially available glucometers (Glucometer M+, Companion 2, HemoCue, One Touch II, and ExacTech Pen) in the hyperbaric environment. WBG samples were prepared, ranging from 25 to 250 mg/dl, for testing glucometers at ground level and at 2.36 atmospheres absolute (ATA). It was noted that at 2.36 ATA, glucose values increased using the Glucometer M+, but decreased with the Companion 2 and HemoCue. The One Touch II values decreased in the hyperglycemic ranges (> 150 mg/dl), whereas the ExacTech Pen monitor readings increased in the hypoglycemic ranges (< 100 mg/dl). The accuracy of WBG monitors is significantly affected by changes in atmospheric pressure.
Atmospheric Pressure↗
Laser-excited fluorescence detection of gas-phase chromatography eluates.
A high-spectral-resolution laser-excited molecular fluorescence gas chromatographic (GC) detection system is evaluated. It utilizes a pulsed supersonic jet expansion to yield very-narrow-bandwidth (e.g. less than or equal to 0.1 nm) fluorescence excitation spectra that may be rapidly scanned via a tunable dye laser. A microcomputer synchronizes the entire system, collects data, performs calculations, and creates visual displays of results. The laser-excited fluorescence detection system was interfaced to a gas chromatograph to exploit both the selectivity of the chromatography and the excellent spectral selectivity of the detector. Fluorescence excitation chromatograms were acquired by monitoring fluorescence emission from selected transition wavelengths characteristic of the GC eluates. The excitation wavelengths were also programmed to change at appropriate retention times to provide greatest selectivities for individual analytes as they eluted, and to allow multiple analytes to be determined in a single elution. Response factors for the system varied appreciably from run to run, which precluded the use of external standard quantitation procedures. However, excellent stabilities for within-run relative response factors were sufficient to allow for good quantitative measurements using internal standard techniques.
Chromatography, Gas↗
Ultraviolet spectrometric detection for gas chromatography of polynuclear aromatic compounds with repetitive spectral scans using absorbance and concurrent fluorescence measurements.
A computer-controlled ultraviolet spectrometric detection system for gas chromatography (GC) is described which acquires absorption spectra of gas-phase GC effluents at rates up to two 62-nm scans per s. Concurrent fluorescence measurements enhance the selectivity of this detection system, providing additional qualitative information. The system has been designed to preserve chromatographic resolution and is selective for aromatic compounds. This multi-mode GC detection is demonstrated through GC separations of polynuclear aromatic compounds with limits of detection ranging from 30 to 70 ng for absorbance measurements. The spectral information obtained with this detection system may be used for compound identification, enhancement of the selectivity and for resolving co-eluting GC eluites.
Chromatography, Gas↗