Serum 1,3-Beta-D-Glucan Values During and After Laparoscopic and Open Intestinal Surgery.

Background1,3-beta-D Glucan (BDG) assay has good accuracy for distinguishing patients with invasive fungal infections from patients without. Some procedures and medications affect BDG levels, resulting in false-positive BDG results. The extent of intestinal surgery on BDG kinetics is unknown. We evaluated the influence of laparoscopic and open intestinal surgery on peri- and postsurgical serum BDG values.MethodsBDG was determined in 346 samples from 50 patients undergoing laparoscopic (24) or open (26) intestinal surgery at the following time points: after insertion of arterial but before skin incision, after skin incision but before dissection of the intestinal mucosa, after completion of anastomosis, after completion of skin sutures, in the evening after surgery, day 2 after surgery, 4-5 days after surgery.ResultsBDG was positive (ie, concentration ≥80 pg/mL) in 54% to 61% of patients during laparoscopic and open surgery (highest rates after completion of skin sutures). BDG was still positive in 12% (open) to 17% (laparoscopic) of patients without any suspected or proven fungal infection or anastomotic leakage 4-5 days after surgery. After completion of gut anastomosis, the BDG increase was higher in open compared with laparoscopic intestinal surgery.ConclusionsThe value of positive BDG tests in the perioperative setting up to 5 days postsurgery seems to be limited due to BDG elevations from intestinal surgical procedures

Post-Operative Functional Outcomes in Early Age Onset Rectal Cancer

Background: Impairment of bowel, urogenital and fertility-related function in patients treated for rectal cancer is common. While the rate of rectal cancer in the young (<50 years) is rising, there is little data on functional outcomes in this group. Methods: The REACCT international collaborative database was reviewed and data on eligible patients analysed. Inclusion criteria comprised patients with a histologically confirmed rectal cancer, <50 years of age at time of diagnosis and with documented follow-up including functional outcomes. Results: A total of 1428 (n=1428) patients met the eligibility criteria and were included in the final analysis. Metastatic disease was present at diagnosis in 13%. Of these, 40% received neoadjuvant therapy and 50% adjuvant chemotherapy. The incidence of post-operative major morbidity was 10%. A defunctioning stoma was placed for 621 patients (43%); 534 of these proceeded to elective restoration of bowel continuity. The median follow-up time was 42 months. Of this cohort, a total of 415 (29%) reported persistent impairment of functional outcomes, the most frequent of which was bowel dysfunction (16%), followed by bladder dysfunction (7%), sexual dysfunction (4.5%) and infertility (1%). Conclusion: A substantial proportion of patients with early-onset rectal cancer who undergo surgery report persistent impairment of functional status. Patients should be involved in the discussion regarding their treatment options and potential impact on quality of life. Functional outcomes should be routinely recorded as part of follow up alongside oncological parameters

Comparison of a Robotic and Patient-Mounted Device for CT-Guided Needle Placement: A Phantom Study

Background: Robotic-based guidance systems are becoming increasingly capable of assisting in needle placement during interventional procedures. Despite these technical advances, less sophisticated low-cost guidance devices promise to enhance puncture accuracy compared with the traditional freehand technique. Purpose: To compare the in vitro accuracy and feasibility of two different aiming devices for computed-tomography (CT)-guided punctures. Methods: A total of 560 CT-guided punctures were performed by using either a robotic (Perfint Healthcare: Maxio) or a novel low-cost patient-mounted system (Medical Templates AG: Puncture Cube System [PCS]) for the placement of Kirschner wires in a plexiglass phantom with different slice thicknesses. Needle placement accuracy as well as procedural time were assessed. The Euclidean (ED) and normal distances (ND) were calculated at the entry and target point. Results: Using the robotic device, the ND at the target for 1.25 mm, 2.5 mm, 3.75 mm and 5 mm slice thickness were 1.28 mm (SD ± 0.79), 1.25 mm (SD ± 0.81), 1.35 mm (SD ± 1.00) and 1.35 mm (SD ± 1.03). Using the PCS, the ND at the target for 1 mm, 3 mm and 5 mm slices were 3.84 mm (SD ± 1.75), 4.41 mm (SD ± 2.31) and 4.41 mm (SD ± 2.11), respectively. With all comparable slice thicknesses, the robotic device was significantly more accurate compared to the low-cost device (p < 0.001). Needle placement with the PCS resulted in lower intervention time (mean, 158.83 s [SD ± 23.38] vs. 225.67 s [SD ± 17.2]). Conclusion: Although the robotic device provided more accurate results, both guidance systems showed acceptable results and may be helpful for interventions in difficult anatomical regions and for those requiring complex multi-angle trajectories

Lentivector-mediated RNAi efficiently suppresses prion protein and prolongs survival of scrapie-infected mice

Prion diseases are fatal neurodegenerative diseases characterized by the accumulation of PrP(Sc), the infectious and protease-resistant form of the cellular prion protein (PrP(C)). We generated lentivectors expressing PrP(C)-specific short hairpin RNAs (shRNAs) that efficiently silenced expression of the prion protein gene (Prnp) in primary neuronal cells. Treatment of scrapie-infected neuronal cells with these lentivectors resulted in an efficient and stable suppression of PrP(Sc) accumulation. After intracranial injection, lentiviral shRNA reduced PrP(C) expression in transgenic mice carrying multiple copies of Prnp. To test the therapeutic potential of lentiviral shRNA, we used what we believe to be a novel approach in which the clinical situation was mimicked. We generated chimeric mice derived from lentivector-transduced embryonic stem cells. Depending on the degree of chimerism, these animals carried the lentiviral shRNAs in a certain percentage of brain cells and expressed reduced levels of PrP(C). Importantly, in highly chimeric mice, survival after scrapie infection was significantly extended. Taken together, these data suggest that lentivector-mediated RNA interference could be an approach for the treatment of prion disease

Substitutions of PrP N-terminal histidine residues modulate scrapie disease pathogenesis and incubation time in transgenic mice.

Prion diseases have been linked to impaired copper homeostasis and copper induced-oxidative damage to the brain. Divalent metal ions, such as Cu2+ and Zn2+, bind to cellular prion protein (PrPC) at octapeptide repeat (OR) and non-OR sites within the N-terminal half of the protein but information on the impact of such binding on conversion to the misfolded isoform often derives from studies using either OR and non-OR peptides or bacterially-expressed recombinant PrP. Here we created new transgenic mouse lines expressing PrP with disrupted copper binding sites within all four histidine-containing OR's (sites 1-4, H60G, H68G, H76G, H84G, "TetraH>G" allele) or at site 5 (composed of residues His-95 and His-110; "H95G" allele) and monitored the formation of misfolded PrP in vivo. Novel transgenic mice expressing PrP(TetraH>G) at levels comparable to wild-type (wt) controls were susceptible to mouse-adapted scrapie strain RML but showed significantly prolonged incubation times. In contrast, amino acid replacement at residue 95 accelerated disease progression in corresponding PrP(H95G) mice. Neuropathological lesions in terminally ill transgenic mice were similar to scrapie-infected wt controls, but less severe. The pattern of PrPSc deposition, however, was not synaptic as seen in wt animals, but instead dense globular plaque-like accumulations of PrPSc in TgPrP(TetraH>G) mice and diffuse PrPSc deposition in (TgPrP(H95G) mice), were observed throughout all brain sections. We conclude that OR and site 5 histidine substitutions have divergent phenotypic impacts and that cis interactions between the OR region and the site 5 region modulate pathogenic outcomes by affecting the PrP globular domain

PrP^C levels and glycosylation profile in healthy wild-type and transgenic mice expressing mutant PrP.

<p><b>(A)</b> Brain homogenates (10 μg per lane) derived from mice expressing full length mouse wild-type (wt), PrP(H95G) lines 4, 11 and 13, PrP(TetraH>G) line 34 and PrP null controls (<i>Prnp</i>^0/0) were subjected to immunoblot analysis using monoclonal antibody SHA31. Blots were reprobed for β-actin to control for equal loading. Molecular weight is indicated on the left (in kDa). (<b>B</b>) Removal of N-linked glycans on PrP^C encoded by the PrP(H95G), lines 4, 11, and 13 as well as PrP(TetraH>G), line 34 using peptide N-glycosidase F (PNGase F) and probing of the treated samples with monoclonal antibody SHA31. fl = full-length PrP.</p

Conformational changes measured by <i>in vitro</i> conversion reactions.

<p><b>(A)</b><i>In vitro</i> conversion reactions have been performed with radiolabeled wild-type and PrP^C(TetraH>G) purified from RK13 cells and PrP^Sc purified from brains of RML-infected Tga20 mice as described [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0188989#pone.0188989.ref004" target="_blank">4</a>]. Samples were analyzed by SDS-PAGE-fluorography, and relative conversion efficiencies (CVE) were calculated from band intensities before and after digestion with proteinase K using the formula CVE [%] = [I°_+PK / (I°_-PK*10)]*100. PrP^C with substituted OR histidines (PrP^C(TetraH>G)) is only half as efficient in converting to the misfolded, PK-resistant conformer than wt PrP^C. Mean values ± standard error (SEM) were determined from 11 independent experiments for each PrP^C type. P-values (p (two sided) = 0.07, p (one sided) = 0.036) were obtained by T-Test calculation. (<b>B</b>) Control reactions performed in the absence of PrP^Sc seed.</p

Western blot analysis of brain lysates from RML-infected wt and transgenic mice for total PrP^C and PrP^Sc using monoclonal antibody 4H11.

<p>(<b>A</b>) Brain homogenates from terminally ill wt and PrP(TetraH>G) line 34 mice were either left untreated (- PK) or subjected to digestion with proteinase K (+ PK). Blots were reprobed for β-actin to control for equal loading. Bands corresponding to total PrP are marked on the left. Irrelevant lanes have been excised at two positions. Molecular weight standards are given on the right (in kDa). (<b>B</b>) Corresponding immunoblot analysis of brain homogenates extracted from RML-infected PrP(H95G) mice from the three different lines 11, 13, and 4, respectively, and corresponding wt control (lanes 1 and 2) before (-) and after (+) treatment with PK. Molecular weight standards are given on the right (in kDa).</p