Tight contact technique during side-to-side laser tissue soldering of rabbit aortas improves tensile strength

BACKGROUND: Cerebral revascularization may be indicated either for blood flow preservation or flow augmentation, often in clinical situations where neither endovascular nor standard surgical intervention can be performed. Cerebral revascularization can be performed by using a temporary occlusive or a non-occlusive technique. Both of these possibilities have their specific range of feasibility. Therefore non-occlusive revascularization techniques have been developed. To further reduce the risks for patients, less time consuming, sutureless techniques such as laser tissue soldering are currently being investigated. METHOD: In the present study, a new technique for side-to-side anastomosis was developed. Using a "sandwich technique", two vessels are kept in close contact during the laser soldering. Thoraco-abdominal aortas from 24 different rabbits were analyzed for laser irradiation induced tensile strength. Two different irradiation modes (continuous and pulsed) were used. The results were compared to conventional, noncontact laser soldering. Histology was performed using HE, Mason's Trichrome staining. FINDINGS: The achieved tensile strengths were significantly higher using the close contact "sandwich technique" as compared to the conventional adaptation technique. Furthermore, tensile strength was higher in the continuously irradiated specimen as compared to the specimen undergoing pulsed laser irradiation. The histology showed similar denaturation areas in both groups. The addition of a collagen membrane between vessel components reduced the tensile strength. CONCLUSION: These first results proved the importance of close and tight contact during the laser soldering procedure thus enabling the development of a "sandwich laser irradiation device" for in vivo application in the rabbit.

GABAergic modulation in central sensitization in humans: a randomized placebo-controlled pharmacokinetic-pharmacodynamic study comparing clobazam with clonazepam in healthy volunteers

Positive allosteric modulators of GABAA receptors (GAMs) acting at specific subtypes of GABAA receptors effectively restore compromised spinal pain control in rodents. Studies addressing a similar antihyperalgesic effect in humans are sparse and are hampered by sedative effects of nonselective GAMs available for use in humans. We present results from a randomized controlled double-blind crossover study in 25 healthy volunteers, which addressed potential antihyperalgesic actions of clobazam (CBZ) and clonazepam (CLN) at mildly sedating equianticonvulsive doses. Clobazam was chosen because of its relatively low sedative properties and CLN because of its use in neuropathic pain. Tolterodine (TLT) was used as an active placebo. The primary outcome parameter was a change in the area of cutaneous UVB irradiation-induced secondary hyperalgesia (ASH), which was monitored for 8 hours after drug application. Sedative effects were assessed in parallel to antihyperalgesia. Compared with TLT, recovery from hyperalgesia was significantly faster in the CBZ and CLN groups (P = 0.009). At the time point of maximum effect, the rate of recovery from hyperalgesia was accelerated by CBZ and CLN, relative to placebo by 15.7% (95% confidence interval [CI] 0.8-30.5), P = 0.040, and 28.6% (95% CI 4.5-52.6), P = 0.022, respectively. Active compounds induced stronger sedation than placebo, but these differences disappeared 8 hours after drug application. We demonstrate here that GAMs effectively reduce central sensitization in healthy volunteers. These results provide proof-of-principle evidence supporting efficacy of GAMs as antihyperalgesic agents in humans and should stimulate further research on compounds with improved subtype specificity.

Proteomic changes in the rat brain induced by homogenous irradiation and by the bystander effect resulting from high energy synchrotron X-ray microbeams

Abstract Purpose: To further evaluate the use of microbeam irradiation (MBI) as a potential means of non-invasive brain tumor treatment by investigating the induction of a bystander effect in non-irradiated tissue. Methods: Adult rats were irradiated with 35 or 350 Gy at the European Synchotron Research Facility (ESRF), using homogenous (broad beam) irradiation (HI) or a high energy microbeam delivered to the right brain hemisphere only. The proteome of the frontal lobes were then analyzed using two-dimensional electrophoresis (2-DE) and mass spectrometry. Results: HI resulted in proteomic responses indicative of tumourigenesis; increased albumin, aconitase and triosphosphate isomerase (TPI), and decreased dihydrolipoyldehydrogenase (DLD). The MBI bystander effect proteomic changes were indicative of reactive oxygen species mediated apoptosis; reduced TPI, prohibitin and tubulin and increased glial fibrillary acidic protein (GFAP). These potentially anti-tumourigenic apoptotic proteomic changes are also associated with neurodegeneration. However the bystander effect also increased heat shock protein (HSP) 71 turnover. HSP 71 is known to protect against all of the neurological disorders characterized by the bystander effect proteome changes. Conclusions: These results indicate that the collective interaction of these MBI-induced bystander effect proteins and their mediation by HSP 71, may confer a protective effect which now warrants additional experimental attention.

Effect of laser soldering irradiation on covalent bonds of pure collagen

Laser tissue welding and soldering is being increasingly used in the clinical setting for defined surgical procedures. The exact induced changes responsible for tensile strength are not yet fully investigated. To further improve the strength of the bonding, a better understanding of the laser impact at the subcellular level is necessary. The goal of this study was to analyze whether the effect of laser irradiation on covalent bonding in pure collagen using irradiances typically applied for tissue soldering. Pure rabbit and equine type I collagen were subjected to laser irradiation. In the first part of the study, rabbit and equine collagen were compared using identical laser and irradiation settings. In the second part of the study, equine collagen was irradiated at increasing laser powers. Changes in covalent bonding were studied indirectly using the sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) technique. Tensile strengths of soldered membranes were measured with a calibrated tensile force gauge. In the first experiment, no differences between the species-specific collagen bands were noted, and no changes in banding were found on SDS-PAGE after laser irradiation. In the second experiment, increasing laser irradiation power showed no effect on collagen banding in SDS-PAGE. Finally, the laser tissue soldering of pure collagen membranes showed virtually no determinable tensile strength. Laser irradiation of pure collagen at typical power settings and exposure times generally used in laser tissue soldering does not induce covalent bonding between collagen molecules. This is true for both rabbit and equine collagen proveniences. Furthermore, soldering of pure collagen membranes without additional cellular components does not achieve the typical tensile strength reported in native, cell-rich tissues. This study is a first step in a better understanding of laser impact at the molecular level and might prove useful in engineering of combined collagen-soldering matrix membranes for special laser soldering applications.

Synchrotron x ray induced axonal transections in the brain of rats assessed by high-field diffusion tensor imaging tractography

Since approximately two thirds of epileptic patients are non-eligible for surgery, local axonal fiber transections might be of particular interest for them. Micrometer to millimeter wide synchrotron-generated X-ray beamlets produced by spatial fractionation of the main beam could generate such fiber disruptions non-invasively. The aim of this work was to optimize irradiation parameters for the induction of fiber transections in the rat brain white matter by exposure to such beamlets. For this purpose, we irradiated cortex and external capsule of normal rats in the antero-posterior direction with a 4 mm×4 mm array of 25 to 1000 µm wide beamlets and entrance doses of 150 Gy to 500 Gy. Axonal fiber responses were assessed with diffusion tensor imaging and fiber tractography; myelin fibers were examined histopathologically. Our study suggests that high radiation doses (500 Gy) are required to interrupt axons and myelin sheaths. However, a radiation dose of 500 Gy delivered by wide minibeams (1000 µm) induced macroscopic brain damage, depicted by a massive loss of matter in fiber tractography maps. With the same radiation dose, the damage induced by thinner microbeams (50 to 100 µm) was limited to their paths. No macroscopic necrosis was observed in the irradiated target while overt transections of myelin were detected histopathologically. Diffusivity values were found to be significantly reduced. A radiation dose ≤ 500 Gy associated with a beamlet size of < 50 µm did not cause visible transections, neither on diffusion maps nor on sections stained for myelin. We conclude that a peak dose of 500 Gy combined with a microbeam width of 100 µm optimally induced axonal transections in the white matter of the brain.

The Drosophila chk2 gene loki is essential for embryonic DNA double-strand-break checkpoints induced in S phase or G2

Cell cycle checkpoints are signal transduction pathways that control the order and timing of cell cycle transitions, ensuring that critical events are completed before the occurrence of the next cell cycle transition. The Chk2 family of kinases is known to play a central role in mediating the cellular responses to DNA damage or DNA replication blocks in various organisms. Here we show through a phylogenetic study that the Drosophila melanogaster serine/threonine kinase Loki is the homolog of the yeast Mek1p, Rad53p, Dun1p, and Cds1 proteins as well as the human Chk2. Functional analyses allowed us to conclude that, in flies, chk2 is involved in monitoring double-strand breaks (DSBs) caused by irradiation during S and G2 phases. In this process it plays an essential role in inducing a cell cycle arrest in embryonic cells. Our results also show that, in contrast to C. elegans chk2, Drosophila chk2 is not essential for normal meiosis and recombination, and it also appears to be dispensable for the MMS-induced DNA damage checkpoint and the HU-induced DNA replication checkpoint during larval development. In addition, Drosophila chk2 does not act at the same cell cycle phases as its yeast homologs, but seems rather to be involved in a pathway similar to the mammalian one, which involves signaling through the ATM/Chk2 pathway in response to genotoxic insults. As mutations in human chk2 were linked to several cancers, these similarities point to the usefulness of the Drosophila model system.

An Intergenic Regulatory Region Mediates Drosophila Myc -Induced Apoptosis and Blocks Tissue Hyperplasia

Induction of cell-autonomous apoptosis following oncogene-induced overproliferation is a major tumor-suppressive mechanism in vertebrates. However, the detailed mechanism mediating this process remains enigmatic. In this study, we demonstrate that dMyc-induced cell-autonomous apoptosis in the fruit fly Drosophila melanogaster relies on an intergenic sequence termed the IRER (irradiation-responsive enhancer region). The IRER mediates the expression of surrounding proapoptotic genes, and we use an in vivo reporter of the IRER chromatin state to gather evidence that epigenetic control of DNA accessibility within the IRER is an important determinant of the strength of this response to excess dMyc. In a previous work, we showed that the IRER also mediates P53-dependent induction of proapoptotic genes following DNA damage, and the chromatin conformation within IRER is regulated by polycomb group-mediated histone modifications. dMyc-induced apoptosis and the P53-mediated DNA damage response thus overlap in a requirement for the IRER. The epigenetic mechanisms controlling IRER accessibility appear to set thresholds for the P53- and dMyc-induced expression of apoptotic genes in vivo and may have a profound impact on cellular sensitivity to oncogene-induced stress.

Study of the radioactivity induced in air by a 15-MeV proton beam.

Radioactivity induced by a 15-MeV proton beam extracted into air was studied at the beam transport line of the 18-MeV cyclotron at the Bern University Hospital (Inselspital). The produced radioactivity was calculated and measured by means of proportional counters located at the main exhaust of the laboratory. These devices were designed for precise assessment of air contamination for radiation protection purposes. The main produced isotopes were 11C, 13N and 14O. Both measurements and calculations correspond to two different irradiation conditions. In the former, protons were allowed to travel for their full range in air. In the latter, they were stopped at the distance of 1.5 m by a beam dump. Radioactivity was measured continuously in the exhausted air starting from 2 min after the end of irradiation. For this reason, the short-lived 14O isotope gave a negligible contribution to the measured activity. Good agreement was found between the measurements and the calculations within the estimated uncertainties. Currents in the range of 120–370 nA were extracted in air for 10–30 s producing activities of 9–22 MBq of 11C and 13N. The total activities for 11C and 13N per beam current and irradiation time for the former and the latter irradiation conditions were measured to be (3.60 ± 0.48) × 10−3 MBq (nA s)−1 and (2.89 ± 0.37) × 10−3 MBq (nA s)−1, respectively.