DNA-lipid systems: A physical chemistry study

It is well known that the interaction of polyelectrolytes with oppositely charged surfactants leads to an associative phase separation; however, the phase behavior of DNA and oppositely charged surfactants is more strongly associative than observed in other systems. A precipitate is formed with very low amounts of surfactant and DNA. DNA compaction is a general phenomenon in the presence of multivalent ions and positively charged surfaces; because of the high charge density there are strong attractive ion correlation effects. Techniques like phase diagram determinations, fluorescence microscopy, and ellipsometry were used to study these systems. The interaction between DNA and catanionic mixtures (i.e., mixtures of cationic and anionic surfactants) was also investigated. We observed that DNA compacts and adsorbs onto the surface of positively charged vesicles, and that the addition of an anionic surfactant can release DNA back into solution from a compact globular complex between DNA and the cationic surfactant. Finally, DNA interactions with polycations, chitosans with different chain lengths, were studied by fluorescence microscopy, in vivo transfection assays and cryogenic transmission electron microscopy. The general conclusion is that a chitosan effective in promoting compaction is also efficient in transfection.

Mechanism of the transforming growth factor-β induction of fibronectin expression in hepatic stem-like cells

Transforming growth factor-β1 (TGF-β1) plays an important role in the fibrogenic process in the liver. The aim of the present study was to explore the action of TGF-β1 on fibronectin expression in rat hepatic stem-like cells and the underlying mechanisms. The level of fibronectin expression was determined in hepatic stem-like cells (WB cells) before and after TGF-β1 stimulation by RT-PCR and Western blot methods. Using immunogold transmission electron microscopy and the Western blot method, we observed the result of the expression and the distribution of cAMP, phosphorylated Smad3 and Smad7 before and after TGF-β1 treatment. The levels of fibronectin expression in both mRNA and protein increased 4- to 5-fold after TGF-β1 stimulation, reaching an optimum level after 8 h and then gradually falling back. Similarly, TGF-β1 stimulation resulted in an increase of cAMP in WB cells, peaking at 8 h. After treatment with TGF-β1 for 24 h, the expression of cAMP gradually decreased. In addition, we found that TGF-β1 treatment also contributed to the increased expression and to changes in cellular distribution of phosphorylated Smad3 (translocation from the cytoplasm to the nucleus) and Smad7 (translocation from the nucleus to the cytoplasm) in WB cells. The present study demonstrates that TGF-β is involved in the fibrogenic process in hepatic stem cells through up-regulation of fibronectin expression, and the mechanisms underlying this process may be associated with the activation of cAMP and Smad pathways.

Electron paramagnetic resonance study of lipid and protein membrane components of erythrocytes oxidized with hydrogen peroxide

Electron paramagnetic resonance (EPR) spectroscopy of spin labels was used to monitor membrane dynamic changes in erythrocytes subjected to oxidative stress with hydrogen peroxide (H2O2). The lipid spin label, 5-doxyl stearic acid, responded to dramatic reductions in membrane fluidity, which was correlated with increases in the protein content of the membrane. Membrane rigidity, associated with the binding of hemoglobin (Hb) to the erythrocyte membrane, was also indicated by a spin-labeled maleimide, 5-MSL, covalently bound to the sulfhydryl groups of membrane proteins. At 2% hematocrit, these alterations in membrane occurred at very low concentrations of H2O2 (50 µM) after only 5 min of incubation at 37°C in azide phosphate buffer, pH 7.4. Lipid peroxidation, suggested by oxidative hemolysis and malondialdehyde formation, started at 300 µM H2O2 (for incubation of 3 h), which is a concentration about six times higher than those detected with the probes. Ascorbic acid and α-tocopherol protected the membrane against lipoperoxidation, but did not prevent the binding of proteins to the erythrocyte membrane. Moreover, the antioxidant (+)-catechin, which also failed to prevent the cross-linking of cytoskeletal proteins with Hb, was very effective in protecting erythrocyte ghosts from lipid peroxidation induced by the Fenton reaction. This study also showed that EPR spectroscopy can be useful to assess the molecular dynamics of red blood cell membranes in both the lipid and protein domains and examine oxidation processes in a system that is so vulnerable to oxidation.

Accuracy of dose planning for prostate radiotherapy in the presence of metallic implants evaluated by electron spin resonance dosimetry

Radiotherapy is one of the main approaches to cure prostate cancer, and its success depends on the accuracy of dose planning. A complicating factor is the presence of a metallic prosthesis in the femur and pelvis, which is becoming more common in elderly populations. The goal of this work was to perform dose measurements to check the accuracy of radiotherapy treatment planning under these complicated conditions. To accomplish this, a scale phantom of an adult pelvic region was used with alanine dosimeters inserted in the prostate region. This phantom was irradiated according to the planned treatment under the following three conditions: with two metallic prostheses in the region of the femur head, with only one prosthesis, and without any prostheses. The combined relative standard uncertainty of dose measurement by electron spin resonance (ESR)/alanine was 5.05%, whereas the combined relative standard uncertainty of the applied dose was 3.35%, resulting in a combined relative standard uncertainty of the whole process of 6.06%. The ESR dosimetry indicated that there was no difference (P>0.05, ANOVA) in dosage between the planned dose and treatments. The results are in the range of the planned dose, within the combined relative uncertainty, demonstrating that the treatment-planning system compensates for the effects caused by the presence of femur and hip metal prostheses.

A dose-response curve for biodosimetry from a 6 MV electron linear accelerator

Biological dosimetry (biodosimetry) is based on the investigation of radiation-induced biological effects (biomarkers), mainly dicentric chromosomes, in order to correlate them with radiation dose. To interpret the dicentric score in terms of absorbed dose, a calibration curve is needed. Each curve should be constructed with respect to basic physical parameters, such as the type of ionizing radiation characterized by low or high linear energy transfer (LET) and dose rate. This study was designed to obtain dose calibration curves by scoring of dicentric chromosomes in peripheral blood lymphocytes irradiated in vitro with a 6 MV electron linear accelerator (Mevatron M, Siemens, USA). Two software programs, CABAS (Chromosomal Aberration Calculation Software) and Dose Estimate, were used to generate the curve. The two software programs are discussed; the results obtained were compared with each other and with other published low LET radiation curves. Both software programs resulted in identical linear and quadratic terms for the curve presented here, which was in good agreement with published curves for similar radiation quality and dose rates.

Effects of intracutaneous injections of sterile water in patients with acute low back pain: a randomized, controlled, clinical trial

Intracutaneous sterile water injection (ISWI) is used for relief of low back pain during labor, acute attacks of urolithiasis, chronic neck and shoulder pain following whiplash injuries, and chronic myofascial pain syndrome. We conducted a randomized, double-blinded, placebo-controlled trial to evaluate the effect of ISWI for relief of acute low back pain (aLBP). A total of 68 patients (41 females and 27 males) between 18 and 55 years old experiencing aLBP with moderate to severe pain (scores ≥5 on an 11-point visual analogue scale [VAS]) were recruited and randomly assigned to receive either ISWIs (n=34) or intracutaneous isotonic saline injections (placebo treatment; n=34). The primary outcome was improvement in pain intensity using the VAS at 10, 45, and 90 min and 1 day after treatment. The secondary outcome was functional improvement, which was assessed using the Patient-Specific Functional Scale (PSFS) 1 day after treatment. The mean VAS score was significantly lower in the ISWI group than in the control group at 10, 45, and 90 min, and 1 day after injection (P<0.05, t-test). The mean increment in PSFS score of the ISWI group was 2.9±2.2 1 day after treatment, while that in the control group was 0.9±2.2. Our study showed that ISWI was effective for relieving pain and improving function in aLBP patients at short-term follow-up. ISWI might be an alternative treatment for aLBP patients, especially in areas where medications are not available, as well as in specific patients (e.g., those who are pregnant or have asthma), who are unable to receive medications or other forms of analgesia because of side effects.

Effect of the addition of wheat fiber and partial pork back fat on the chemical composition, texture and sensory property of low-fat bologna sausage containing inulin and oat fiber

The objective of this work was to study the effect of adding wheat fiber and partial pork back fat on the quality characteristics of bologna sausage. The compound central rotating design was used with treatments containing fixed levels of inulin (5%) and oat fiber (1%) and variable levels of wheat fiber (0-4%) and pork back fat (0-10%). The pH and protein were similar in all the treatments, the fat was lower than the control treatment and the moisture content was higher than the control treatment (CF) without fibers. The wheat fiber increased the hardness and reduced cohesiveness and scores were given for overall impression. We found that it was possible to prepare low-fat bologna sausage with the addition of 6.58% fiber (5% inulin, 1% oat fiber and 0.58% wheat fiber), whilst retaining good sensory acceptability, thus reducing the pork back fat levels by between 25 and 42.75%.