[HbA1c revisited (again and again ...)]

The "International Federation of Clinical Chemistry" (IFCC) has developed a new international reference measurement system as anchor for worldwide standardization of HbA1c determinations. The use of IFCC-referenced methods results in "true" HbA1c-values that are 1 - 2% lower compared to traditional methods. This leads to potential risks of clinical misjudgement. For the evaluation of glycaemic control it is, therefore, important to know the method used, its normal range as well as possibilities of mathematical conversion of results. To reduce the risk of wrong interpretation of results the Swiss Society of Endocrinology and Diabetes recommends that reports of HbA1c clearly indicate whether the values are "IFCC-" or "DCCT-traceable". "IFCC"-results should best be reported in mmol/mol. In addition, the normal range has to be indicated properly.

Intracellular localization of the BCL-2 family member BOK and functional implications

The pro-apoptotic BCL-2 family member BOK is widely expressed and resembles the multi-BH domain proteins BAX and BAK based on its amino acid sequence. The genomic region encoding BOK was reported to be frequently deleted in human cancer and it has therefore been hypothesized that BOK functions as a tumor suppressor. However, little is known about the molecular functions of BOK. We show that enforced expression of BOK activates the intrinsic (mitochondrial) apoptotic pathway in BAX/BAK-proficient cells but fails to kill cells lacking both BAX and BAK or sensitize them to cytotoxic insults. Interestingly, major portions of endogenous BOK are localized to and partially inserted into the membranes of the Golgi apparatus as well as the endoplasmic reticulum (ER) and associated membranes. The C-terminal transmembrane domain of BOK thereby constitutes a 'tail-anchor' specific for targeting to the Golgi and ER. Overexpression of full-length BOK causes early fragmentation of ER and Golgi compartments. A role for BOK on the Golgi apparatus and the ER is supported by an abnormal response of Bok-deficient cells to the Golgi/ER stressor brefeldin A. Based on these results, we propose that major functions of BOK are exerted at the Golgi and ER membranes and that BOK induces apoptosis in a manner dependent on BAX and BAK.

Single-Molecule Conductance of Functionalized Oligoynes: Length Dependence and Junction Evolution

We report a combined experimental and theoretical investigation of the length dependence and anchor group dependence of the electrical conductance of a series of oligoyne molecular wires in single-molecule junctions with gold contacts. Experimentally, we focus on the synthesis and properties of diaryloligoynes with n = 1, 2, and 4 triple bonds and the anchor dihydrobenzo[b]thiophene (BT). For comparison, we also explored the aurophilic anchor group cyano (CN), amino (NH2), thiol (SH), and 4-pyridyl (PY). Scanning tunneling microscopy break junction (STM-BJ) and mechanically controllable break junction (MCBJ) techniques are employed to investigate single-molecule conductance characteristics. The BT moiety is superior as compared to traditional anchoring groups investigated so far. BT-terminated oligoynes display a 100% probability of junction formation and possess conductance values which are the highest of the oligoynes studied and, moreover, are higher than other conjugated molecular wires of similar length. Density functional theory (DFT)-based calculations are reported for oligoynes with n = 1−4 triple bonds. Complete conductance traces and conductance distributions are computed for each family of molecules. The sliding of the anchor groups leads to oscillations in both the electrical conductance and the binding energies of the studied molecular wires. In agreement with experimental results, BT-terminated oligoynes are predicted to have a high electrical conductance. The experimental attenuation constants βH range between 1.7 nm−1 (CN) and 3.2 nm−1 (SH) and show the following trend: βH(CN) < βH(NH2) < βH(BT) < βH(PY) ≈ βH(SH). DFT-based calculations yield lower values, which range between 0.4 nm−1 (CN) and 2.2 nm−1 (PY).

Balancing on a slackline: 8-year-olds vs. adults

Children are less stable than adults during static upright stance. We investigated whether the same holds true for a task that was novel for both children and adults and highly dynamic: single-legged stance on a slackline. We compared 8-year-olds with young adults and assessed the following outcome measures: time on the slackline, stability on the slack-line (calculated from slackline reaction force), gaze movement, head-in-space rotation and translation, trunk-in-space rotation, and head-on-trunk rotation. Eight-year-olds fell off the slackline quicker and were generally less stable on the slackline than adults. Eight-year-olds also showed more head-in-space rotation and translation, and more gaze variability around a visual anchor point they were instructed to fixate. Trunk-in-space and head-on-trunk rotations did not differ between groups. The results imply that the lower postural stability of 8-year-olds compared to adults – as found in simple upright stance – holds true for dynamic, novel tasks in which adults lack the advantage of more practice. They also suggest that the lack of head and gaze stability constitutes an important limiting factor in children’s ability to master such tasks

Novel prototype sewing device, EndoSew ® , for minimally invasive surgery: an extracorporeal ileal conduit construction pilot study in 10 patients

What's known on the subject? and What does the study add? The EndoSew® prototype was first tested in a porcine model several years ago. The investigators found it both simple to master and reliable, its greatest advantage being a 2.4-fold time saving compared with straight laparoscopic suturing. In addition to that publication, there is a single case report describing the performance of an open EndoSew® suture to close parts (16 cm) of an ileal neobladder. The time for suturing the 16 cm ileum was 25 min, which is in line with our experience. The knowledge on this subject is limited to these two publications. We report on the first consecutive series of ileal conduits performed in humans using the novel prototype sewing device EndoSew®. The study shows that the beginning and the end of the suture process represent the critical procedural steps. It also shows that, overall, the prototype sewing machine has the potential to facilitate the intracorporeal suturing required in reconstructive urology for construction of urinary diversions. Objective To evaluate the feasibility and safety of the novel prototype sewing device EndoSew® in placing an extracorporeal resorbable running suture for ileal conduits. Patients and Methods We conducted a prospective single-centre pilot study of 10 consecutive patients undergoing ileal conduit, in whom the proximal end of the ileal conduit was closed extracorporeally using an EndoSew® running suture. The primary endpoint was the safety of the device and the feasibility of the sewing procedure which was defined as a complete watertight running suture line accomplished by EndoSew® only. Watertightness was assessed using methylene blue intraoperatively and by loopography on postoperative days 7 and 14. Secondary endpoints were the time requirements and complications ≤30 days after surgery. Results A complete EndoSew® running suture was feasible in nine patients; the suture had to be abandoned in one patient because of mechanical failure. In three patients, two additional single freehand stitches were needed to anchor the thread and to seal tiny leaks. Consequently, all suture lines in 6/10 patients were watertight with EndoSew® suturing alone and in 10/10 patients after additional freehand stitches. The median (range) sewing time was 5.5 (3–10) min and the median (range) suture length was 4.5 (2–5.5) cm. There were no suture-related complications. Conclusions The EndoSew® procedure is both feasible and safe. After additional freehand stitches in four patients all sutures were watertight. With further technical refinements, EndoSew® has the potential to facilitate the intracorporeal construction of urinary diversions.

HLA-B⁎27 subtype specificity determines targeting and viral evolution of a hepatitis C virus-specific CD8+ T cell epitope

Background & Aims: HLA-B⁄27 is associated with spontaneous HCV genotype 1 clearance. HLA-B⁄27-restricted CD8+ T cells target three NS5B epitopes. Two of these epitopes are dominantly targeted in the majority of HLA-B⁄27+ patients. In chronic infection, viral escape occurs consistently in these two epitopes. The third epitope (NS5B2820) was dominantly targeted in an acutely infected patient. This was in contrast, however, to the lack of recognition and viral escape in the large majority of HLA-B⁄27+ patients. Here, we set out to determine the host factors contributing to selective targeting of this epitope. Methods: Four-digit HLA class I typing and viral sequence analyses were performed in 78 HLA-B⁄27+ patients with chronic HCV genotype 1 infection. CD8+ T cell analyses were performed in a subset of patients. In addition, HLA/peptide affinity was compared for HLA-B⁄27:02 and 05. Results: The NS5B2820 epitope is only restricted by the HLA-B⁄27 subtype HLA-B⁄27:02 (that is frequent in Mediterranean populations), but not by the prototype HLA-B⁄27 subtype B⁄27:05. Indeed, the epitope is very dominant in HLA-B⁄27:02+ patients and is associated with viral escape mutations at the anchor position for HLA-binding in 12 out of 13 HLA-B⁄27:02+ chronically infected patients. Conclusions: The NS5B2820 epitope is immunodominant in the context of HLA-B⁄27:02, but is not restricted by other HLA-B⁄27 subtypes. This finding suggests an important role of HLA subtypes in the restriction of HCV-specific CD8+ responses. With minor HLA subtypes covering up to 39% of specific populations, these findings may have important implications for the selection of epitopes for global vaccines.

Ezrin/Radixin/Moesin proteins and flotillins cooperate to promote uropod formation in T cells

T cell uropods are enriched in specific proteins including adhesion receptors such as P-selectin glycoprotein ligand-1 (PSGL-1), lipid raft-associated proteins such as flotillins and ezrin/radixin/moesin (ERM) proteins which associate with cholesterol-rich raft domains and anchor adhesion receptors to the actin cytoskeleton. Using dominant mutants and siRNA technology we have tested the interactions among these proteins and their role in shaping the T cell uropod. Expression of wild type (WT) ezrin-EGFP failed to affect the morphology of human T cells or chemokine-induced uropod recruitment of PSGL-1 and flotillin-1 and -2. In contrast, expression of constitutively active T567D ezrin-EGFP induced a motile, polarized phenotype in some of the transfected T cells, even in the absence of chemokine. These cells featured F-actin-rich ruffles in the front and uropod enrichment of PSGL-1 and flotillins. T567D ezrin-EGFP was itself strongly enriched in the rear of the polarized T cells. Uropod formation induced by T567D ezrin-EGFP was actin-dependent as it was attenuated by inhibition of Rho-kinase or myosin II, and abolished by disruption of actin filaments. While expression of constitutively active ezrin enhanced cell polarity, expression of a dominant-negative deletion mutant of ezrin, 1-310 ezrin-EGFP, markedly reduced uropod formation induced by the chemokine SDF-1, T cell front-tail polarity, and capping of PSGL-1 and flotillins. Transfection of T cells with WT or T567D ezrin did not affect chemokine-mediated chemotaxis whereas 1-310 ezrin significantly impaired spontaneous 2D migration and chemotaxis. siRNA-mediated downregulation of flotillins in murine T cells attenuated moesin capping and uropod formation, indicating that ERM proteins and flotillins cooperate in uropod formation. In summary, our results indicate that activated ERM proteins function together with flotillins to promote efficient chemotaxis of T cells by structuring the uropod of migrating T cells.

Extraspinal ossifications after implantation of vertical expandable prosthetic titanium ribs (VEPTRs).

INTRODUCTION Though developed for thoracic insufficiency syndrome, the spinal growth-stimulating potential and the ease of placement of vertical expandable titanium ribs (VEPTRs) has resulted in their widespread use for early-onset spine deformity. Observation of implant-related ossifications warrants further assessment, since they may be detrimental to the function-preserving non-fusion strategy. PATIENTS AND METHODS Radiographs (obtained pre and post index procedure, and at 4-year follow-up) and the records of 65 VEPTR patients from four paediatric spine centres were analysed. Ossifications were classified as type I (at anchor points), type II (along the central part) or type III (re-ossification after thoracostomy). RESULTS The average age at the index procedure was 6.5 years (min 1, max 13.7). The most prevalent spine problem was congenital scoliosis (37) with rib fusions (34), followed by neuromuscular and syndromic deformities (13 and 8, respectively). Idiopathic and secondary scoliosis (e.g. after thoracotomy) were less frequent (3 and 4, respectively). Forty-two of the 65 (65 %) patients showed ossifications, half of which were around the anchors. Forty-five percent (15/33) without pre-existing rib fusions developed a type II ossification along the implant. Re-ossifications of thoracostomies were less frequent (5/34, 15 %). The occurrence of ossifications was not associated with patient-specific factors. CONCLUSIONS Implant-related ossifications around VEPTR are common. In contrast to harmless bone formation around anchors, ossifications around the telescopic part and the rod section are troublesome in view of their possible negative impact on chest cage compliance and spinal mobility. This potential side effect needs to be considered during implant selection, particularly in patients with originally normal thoracic and spinal anatomy.

RNA interference screening identifies a novel role for PCTK1/CDK16 in medulloblastoma with c-Myc amplification.

Medulloblastoma (MB) is the most common malignant brain tumor in children and is associated with a poor outcome. cMYC amplification characterizes a subgroup of MB with very poor prognosis. However, there exist so far no targeted therapies for the subgroup of MB with cMYC amplification. Here we used kinome-wide RNA interference screening to identify novel kinases that may be targeted to inhibit the proliferation of c-Myc-overexpressing MB. The RNAi screen identified a set of 5 genes that could be targeted to selectively impair the proliferation of c-Myc-overexpressing MB cell lines: AKAP12 (A-kinase anchor protein), CSNK1α1 (casein kinase 1, alpha 1), EPHA7 (EPH receptor A7) and PCTK1 (PCTAIRE protein kinase 1). When using RNAi and a pharmacological inhibitor selective for PCTK1, we could show that this kinase plays a crucial role in the proliferation of MB cell lines and the activation of the mammalian target of rapamycin (mTOR) pathway. In addition, pharmacological PCTK1 inhibition reduced the expression levels of c-Myc. Finally, targeting PCTK1 selectively impaired the tumor growth of c-Myc-overexpressing MB cells in vivo. Together our data uncover a novel and crucial role for PCTK1 in the proliferation and survival of MB characterized by cMYC amplification.

A Passive Source Localization System for IEEE 802.15.4 Signal

In this work, we provide a passive location monitoring system for IEEE 802.15.4 signal emitters. The system adopts software defined radio techniques to passively overhear IEEE 802.15.4 packets and to extract power information from baseband signals. In our system, we provide a new model based on the nonlinear regression for ranging. After obtaining distance information, a Weighted Centroid (WC) algorithm is adopted to locate users. In WC, each weight is inversely proportional to the nth power of propagation distance, and the degree n is obtained from some initial measurements. We evaluate our system in a 16m-18m area with complex indoor propagation conditions. We are able to achieve a median error of 2:1m with only 4 anchor nodes.

The Synthesis of Functionalised Diaryltetraynes and Their Transport Properties in Single-Molecule Junctions

The synthesis and characterisation is described of six diaryltetrayne derivatives [Ar-(C[TRIPLE BOND]C)4-Ar] with Ar=4-NO2-C6H4- (NO24), 4-NH(Me)C6H4- (NHMe4), 4-NMe2C6H4- (NMe24), 4-NH2-(2,6-dimethyl)C6H4- (DMeNH24), 5-indolyl (IN4) and 5-benzothienyl (BTh4). X-ray molecular structures are reported for NO24, NHMe4, DMeNH24, IN4 and BTh4. The stability of the tetraynes has been assessed under ambient laboratory conditions (20 °C, daylight and in air): NO24 and BTh4 are stable for at least six months without observable decomposition, whereas NHMe4, NMe24, DMeNH24 and IN4 decompose within a few hours or days. The derivative DMeNH24, with ortho-methyl groups partially shielding the tetrayne backbone, is considerably more stable than the parent compound with Ar=4-NH2C6H4 (NH24). The ability of the stable tetraynes to anchor in Au|molecule|Au junctions is reported. Scanning-tunnelling-microscopy break junction (STM-BJ) and mechanically controllable break junction (MCBJ) techniques are employed to investigate single-molecule conductance characteristics.

Prolonged activity of the pestiviral RNase Erns as an interferon antagonist after uptake by clathrin-mediated endocytosis

The RNase activity of the envelope glycoprotein E(rns) of the pestivirus bovine viral diarrhea virus (BVDV) is required to block type I interferon (IFN) synthesis induced by single-stranded RNA (ssRNA) and double-stranded RNA (dsRNA) in bovine cells. Due to the presence of an unusual membrane anchor at its C terminus, a significant portion of E(rns) is also secreted. In addition, a binding site for cell surface glycosaminoglycans is located within the C-terminal region of E(rns). Here, we show that the activity of soluble E(rns) as an IFN antagonist is not restricted to bovine cells. Extracellularly applied E(rns) protein bound to cell surface glycosaminoglycans and was internalized into the cells within 1 h of incubation by an energy-dependent mechanism that could be blocked by inhibitors of clathrin-dependent endocytosis. E(rns) mutants that lacked the C-terminal membrane anchor retained RNase activity but lost most of their intracellular activity as an IFN antagonist. Surprisingly, once taken up into the cells, E(rns) remained active and blocked dsRNA-induced IFN synthesis for several days. Thus, we propose that E(rns) acts as an enzymatically active decoy receptor that degrades extracellularly added viral RNA mainly in endolysosomal compartments that might otherwise activate intracellular pattern recognition receptors (PRRs) in order to maintain a state of innate immunotolerance. IMPORTANCE The pestiviral RNase E(rns) was previously shown to inhibit viral ssRNA- and dsRNA-induced interferon (IFN) synthesis. However, the localization of E(rns) at or inside the cells, its species specificity, and its mechanism of interaction with cell membranes in order to block the host's innate immune response are still largely unknown. Here, we provide strong evidence that the pestiviral RNase E(rns) is taken up within minutes by clathrin-mediated endocytosis and that this uptake is mostly dependent on the glycosaminoglycan binding site located within the C-terminal end of the protein. Remarkably, the inhibitory activity of E(rns) remains for several days, indicating the very potent and prolonged effect of a viral IFN antagonist. This novel mechanism of an enzymatically active decoy receptor that degrades a major viral pathogen-associated molecular pattern (PAMP) might be required to efficiently maintain innate and, thus, also adaptive immunotolerance, and it might well be relevant beyond the bovine species.

Range-based Weighted-likelihood Particle Filter for RSS-based Indoor Tracking

Attractive business cases in various application fields contribute to the sustained long-term interest in indoor localization and tracking by the research community. Location tracking is generally treated as a dynamic state estimation problem, consisting of two steps: (i) location estimation through measurement, and (ii) location prediction. For the estimation step, one of the most efficient and low-cost solutions is Received Signal Strength (RSS)-based ranging. However, various challenges - unrealistic propagation model, non-line of sight (NLOS), and multipath propagation - are yet to be addressed. Particle filters are a popular choice for dealing with the inherent non-linearities in both location measurements and motion dynamics. While such filters have been successfully applied to accurate, time-based ranging measurements, dealing with the more error-prone RSS based ranging is still challenging. In this work, we address the above issues with a novel, weighted likelihood, bootstrap particle filter for tracking via RSS-based ranging. Our filter weights the individual likelihoods from different anchor nodes exponentially, according to the ranging estimation. We also employ an improved propagation model for more accurate RSS-based ranging, which we suggested in recent work. We implemented and tested our algorithm in a passive localization system with IEEE 802.15.4 signals, showing that our proposed solution largely outperforms a traditional bootstrap particle filter.

Fine-grained Indoor Tracking by Fusing Inertial Sensor and Physical Layer Information in WLANs

Indoor positioning has become an emerging research area because of huge commercial demands for location-based services in indoor environments. Channel State Information (CSI) as a fine-grained physical layer information has been recently proposed to achieve high positioning accuracy by using range-based methods, e.g., trilateration. In this work, we propose to fuse the CSI-based ranges and velocity estimated from inertial sensors by an enhanced particle filter to achieve highly accurate tracking. The algorithm relies on some enhanced ranging methods and further mitigates the remaining ranging errors by a weighting technique. Additionally, we provide an efficient method to estimate the velocity based on inertial sensors. The algorithms are designed in a network-based system, which uses rather cheap commercial devices as anchor nodes. We evaluate our system in a complex environment along three different moving paths. Our proposed tracking method can achieve 1:3m for mean accuracy and 2:2m for 90% accuracy, which is more accurate and stable than pedestrian dead reckoning and range-based positioning.

Passively Track WiFi Users with an Enhanced Particle Filter using Power-based Ranging

Passive positioning systems produce user location information for third-party providers of positioning services. Since the tracked wireless devices do not participate in the positioning process, passive positioning can only rely on simple, measurable radio signal parameters, such as timing or power information. In this work, we provide a passive tracking system for WiFi signals with an enhanced particle filter using fine-grained power-based ranging. Our proposed particle filter provides an improved likelihood function on observation parameters and is equipped with a modified coordinated turn model to address the challenges in a passive positioning system. The anchor nodes for WiFi signal sniffing and target positioning use software defined radio techniques to extract channel state information to mitigate multipath effects. By combining the enhanced particle filter and a set of enhanced ranging methods, our system can track mobile targets with an accuracy of 1.5m for 50% and 2.3m for 90% in a complex indoor environment. Our proposed particle filter significantly outperforms the typical bootstrap particle filter, extended Kalman filter and trilateration algorithms.