Population pharmacokinetics of sevoflurane in conjunction with the AnaConDa: toward target-controlled infusion of volatiles into the breathing system

BACKGROUND: The Anesthetic Conserving Device (AnaConDa) uncouples delivery of a volatile anesthetic (VA) from fresh gas flow (FGF) using a continuous infusion of liquid volatile into a modified heat-moisture exchanger capable of adsorbing VA during expiration and releasing adsorbed VA during inspiration. It combines the simplicity and responsiveness of high FGF with low agent expenditures. We performed in vitro characterization of the device before developing a population pharmacokinetic model for sevoflurane administration with the AnaConDa, and retrospectively testing its performance (internal validation). MATERIALS AND METHODS: Eighteen females and 20 males, aged 31-87, BMI 20-38, were included. The end-tidal concentrations were varied and recorded together with the VA infusion rates into the device, ventilation and demographic data. The concentration-time course of sevoflurane was described using linear differential equations, and the most suitable structural model and typical parameter values were identified. The individual pharmacokinetic parameters were obtained and tested for covariate relationships. Prediction errors were calculated. RESULTS: In vitro studies assessed the contribution of the device to the pharmacokinetic model. In vivo, the sevoflurane concentration-time courses on the patient side of the AnaConDa were adequately described with a two-compartment model. The population median absolute prediction error was 27% (interquartile range 13-45%). CONCLUSION: The predictive performance of the two-compartment model was similar to that of models accepted for TCI administration of intravenous anesthetics, supporting open-loop administration of sevoflurane with the AnaConDa. Further studies will focus on prospective testing and external validation of the model implemented in a target-controlled infusion device.

DAPK2 is a novel E2F1/KLF6 target gene involved in their proapoptotic function

Death-associated protein kinase 2 (DAPK2) belongs to a family of proapoptotic Ca(2+)/calmodulin-regulated serine/threonine kinases. We recently identified DAPK2 as an enhancing factor during granulocytic differentiation. To identify transcriptional DAPK2 regulators, we cloned 2.7 kb of the 5'-flanking region of the DAPK2 gene. We found that E2F1 and Krüppel-like factor 6 (KLF6) strongly activate the DAPK2 promoter. We mapped the E2F1 and KLF6 responsive elements to a GC-rich region 5' of exon 1 containing several binding sites for KLF6 and Sp1 but not for E2F. Moreover, we showed that transcriptional activation of DAPK2 by E2F1 and KLF6 is dependent on Sp1 using Sp1/KLF6-deficient insect cells, mithramycin A treatment to block Sp1-binding or Sp1 knockdown cells. Chromatin immunoprecipitation revealed recruitment of Sp1 and to lesser extent that of E2F1 and KLF6 to the DAPK2 promoter. Activation of E2F1 in osteosarcoma cells led to an increase of endogenous DAPK2 paralleled by cell death. Inhibition of DAPK2 expression resulted in significantly reduced cell death upon E2F1 activation. Similarly, KLF6 expression in H1299 cells increased DAPK2 levels accompanied by cell death that is markedly decreased upon DAPK2 knockdown. Moreover, E2F1 and KLF6 show cooperation in activating the DAPK2 promoter. In summary, our findings establish DAPK2 as a novel Sp1-dependent target gene for E2F1 and KLF6 in cell death response.

Nonpeptide somatostatin receptor agonists specifically target ocular neovascularization via the somatostatin type 2 receptor

PURPOSE: To define the molecular pharmacology underlying the antiangiogenic effects of nonpeptide imidazolidine-2,4-dione somatostatin receptor agonists (NISAs) and evaluate the efficacy of NISA in ocular versus systemic delivery routes in ocular disease models. METHODS: Functional inhibitory effects of the NISAs and the somatostatin peptide analogue octreotide were evaluated in vitro by chemotaxis, proliferation, and tube-formation assays. The oxygen-induced retinopathy (OIR) model and the laser model of choroidal neovascularization (CNV) were used to test the in vivo efficacy of NISAs. Transscleral permeability of a candidate NISA was also measured. RESULTS: NISAs inhibited growth factor-induced HREC proliferation, migration and tube formation with submicromolar potencies (IC(50), 0.1-1.0 microM) comparable to octreotide. In the OIR model, systemic administration of the NISAs RFE-007 and RFE-011 inhibited retinal neovascularization in a dose-dependent manner, comparable to octreotide. In the CNV model, intravitreal RFE-011 resulted in a 56% reduction (P < 0.01) in CNV lesion area, whereas systemic administration resulted in a 35% reduction (P < 0.05) in lesion area. RFE-011 demonstrated transscleral penetration. CONCLUSIONS: Micromolar concentrations of octreotide and NISAs are necessary for antiangiogenic effects, whereas nanomolar concentrations are effective for endocrine inhibition. This suggests that the antiangiogenic activity of NISAs and octreotide is mediated by an overall much less efficient downstream coupling mechanism than is growth hormone release. As a result, the intravitreal or transscleral route of administration should be seriously considered for future clinical studies of SSTR2 agonists used for treatment of ocular neovascularization to ensure efficacious concentrations in the target retinal and choroidal tissue.

Natural antibodies target virus-antibody complexes to organized lymphoid tissue

Natural antibodies (NA) specific for infectious pathogens are found at low titer (usually <1:40) in the serum of healthy, non-immunized, individuals. Therefore, NA are part of the first line of defence against blood borne microorganisms. They directly neutralize viral infections or lyse pathogens by activating the complement cascade. In addition, recent studies highlighted their role in the pooling of infectious pathogens and other antigens to the spleen. This prevents infection of vital target organs and enhances the induction of adaptive immune responses. Specific T and B-cell responses are exclusively induced in highly organized secondary lymphoid organs including lymph nodes and the spleen. As a consequence, mice with disrupted microorganisation of lymphoid organs have defective adaptive immunity. In addition, some pathogens including lymphocytic choriomeningitis virus (LCMV), Leishmania and HIV developed strategies to destroy the splenic architecture in order to induce an acquired immunosuppression and to establish persistent infection. NA antibodies enhance early neutralizing antibodies in the absence of T help mainly by targeting antigen to the splenic marginal zone. In addition, by activating the complement cascade, NA enhance T cell and T-cell dependent B-cell responses. Therefore, natural antibodies are an important link between innate and adaptive immunity.

Multi-antenna non-line-of-sight identification techniques for target localization in mobile ad-hoc networks

Target localization has a wide range of military and civilian applications in wireless mobile networks. Examples include battle-field surveillance, emergency 911 (E911), traffc alert, habitat monitoring, resource allocation, routing, and disaster mitigation. Basic localization techniques include time-of-arrival (TOA), direction-of-arrival (DOA) and received-signal strength (RSS) estimation. Techniques that are proposed based on TOA and DOA are very sensitive to the availability of Line-of-sight (LOS) which is the direct path between the transmitter and the receiver. If LOS is not available, TOA and DOA estimation errors create a large localization error. In order to reduce NLOS localization error, NLOS identifcation, mitigation, and localization techniques have been proposed. This research investigates NLOS identifcation for multiple antennas radio systems. The techniques proposed in the literature mainly use one antenna element to enable NLOS identifcation. When a single antenna is utilized, limited features of the wireless channel can be exploited to identify NLOS situations. However, in DOA-based wireless localization systems, multiple antenna elements are available. In addition, multiple antenna technology has been adopted in many widely used wireless systems such as wireless LAN 802.11n and WiMAX 802.16e which are good candidates for localization based services. In this work, the potential of spatial channel information for high performance NLOS identifcation is investigated. Considering narrowband multiple antenna wireless systems, two xvNLOS identifcation techniques are proposed. Here, the implementation of spatial correlation of channel coeffcients across antenna elements as a metric for NLOS identifcation is proposed. In order to obtain the spatial correlation, a new multi-input multi-output (MIMO) channel model based on rough surface theory is proposed. This model can be used to compute the spatial correlation between the antenna pair separated by any distance. In addition, a new NLOS identifcation technique that exploits the statistics of phase difference across two antenna elements is proposed. This technique assumes the phases received across two antenna elements are uncorrelated. This assumption is validated based on the well-known circular and elliptic scattering models. Next, it is proved that the channel Rician K-factor is a function of the phase difference variance. Exploiting Rician K-factor, techniques to identify NLOS scenarios are proposed. Considering wideband multiple antenna wireless systems which use MIMO-orthogonal frequency division multiplexing (OFDM) signaling, space-time-frequency channel correlation is exploited to attain NLOS identifcation in time-varying, frequency-selective and spaceselective radio channels. Novel NLOS identi?cation measures based on space, time and frequency channel correlation are proposed and their performances are evaluated. These measures represent a better NLOS identifcation performance compared to those that only use space, time or frequency.

Tracking a non-cooperative target using a Doppler radar wireless sensor network

Tracking or target localization is used in a wide range of important tasks from knowing when your flight will arrive to ensuring your mail is received on time. Tracking provides the location of resources enabling solutions to complex logistical problems. Wireless Sensor Networks (WSN) create new opportunities when applied to tracking, such as more flexible deployment and real-time information. When radar is used as the sensing element in a tracking WSN better results can be obtained; because radar has a comparatively larger range both in distance and angle to other sensors commonly used in WSNs. This allows for less nodes deployed covering larger areas, saving money. In this report I implement a tracking WSN platform similar to what was developed by Lim, Wang, and Terzis. This consists of several sensor nodes each with a radar, a sink node connected to a host PC, and a Matlab© program to fuse sensor data. I have re-implemented their experiment with my WSN platform for tracking a non-cooperative target to verify their results and also run simulations to compare. The results of these tests are discussed and some future improvements are proposed.

Human basophils and eosinophils are the direct target leukocytes of the novel IL-1 family member IL-33

In mice, interleukin-18 (IL-18) regulates Th1- or Th2-type immune responses depending on the cytokine environment and effector cells involved, and the ST2-ligand, IL-33, primarily promotes an allergic phenotype. Human basophils, major players in allergic inflammation, constitutively express IL-18 receptors, while ST2 surface expression is inducible by IL-3. Unexpectedly, freshly isolated basophils are strongly activated by IL-33, but, in contrast to mouse basophils, do not respond to IL-18. IL-33 promotes IL-4, IL-13 and IL-8 secretion in synergy with IL-3 and/or FcepsilonRI-activation, and enhances FcepsilonRI-induced mediator release. These effects are similar to that of IL-3, but the signaling pathways engaged are distinct because IL-33 strongly activates NF-kappaB and shows a preference for p38 MAP-kinase, while IL-3 acts through Jak/Stat and preferentially activates ERK. Eosinophils are the only other leukocyte-type directly activated by IL-33, as evidenced by screening of p38-activation in peripheral blood cells. Only upon CD3/CD28-ligation, IL-33 weakly enhances Th2 cytokine expression by in vivo polarized Th2 cells. This study on primary human cells demonstrates that basophils and eosinophils are the only direct target leukocytes for IL-33, suggesting that IL-33 promotes allergic inflammation and Th2 polarization mainly by the selective activation of these specialized cells of the innate immune system.

Adaptive sensing for target tracking applications

In a statistical inference scenario, the estimation of target signal or its parameters is done by processing data from informative measurements. The estimation performance can be enhanced if we choose the measurements based on some criteria that help to direct our sensing resources such that the measurements are more informative about the parameter we intend to estimate. While taking multiple measurements, the measurements can be chosen online so that more information could be extracted from the data in each measurement process. This approach fits well in Bayesian inference model often used to produce successive posterior distributions of the associated parameter. We explore the sensor array processing scenario for adaptive sensing of a target parameter. The measurement choice is described by a measurement matrix that multiplies the data vector normally associated with the array signal processing. The adaptive sensing of both static and dynamic system models is done by the online selection of proper measurement matrix over time. For the dynamic system model, the target is assumed to move with some distribution and the prior distribution at each time step is changed. The information gained through adaptive sensing of the moving target is lost due to the relative shift of the target. The adaptive sensing paradigm has many similarities with compressive sensing. We have attempted to reconcile the two approaches by modifying the observation model of adaptive sensing to match the compressive sensing model for the estimation of a sparse vector.

Integrin alphavbeta6 is a marker of the progression of biliary and portal liver fibrosis and a novel target for antifibrotic therapies

BACKGROUND/AIMS: The integrin alphavbeta6 promotes proliferation of specialized epithelia and acts as a receptor for the activation of latent TGFbeta1. We studied alphavbeta6 expression in experimental and human liver fibrosis and the potential of its pharmacological inhibition for treatment of hepatic fibrosis. METHODS: alphavbeta6 expression was studied by quantitative PCR and immunohistochemistry in rats with cirrhosis due to bile duct ligation (BDL), administration of thioacetamide (TAA), in Mdr2(Abcb4)(-/-) mice with spontaneous biliary fibrosis, and in livers of patients with chronic hepatitis C (n=79) and end-stage liver disease due to various etiologies (n=18). The effect of a selective alphavbeta6 inhibitor was evaluated in Mdr2(Abcb4)(-/-) mice with ongoing fibrogenesis. RESULTS: Integrin beta6 mRNA increased with fibrosis stage in hepatitis C and was upregulated between 25- and 100-fold in TAA- and BDL-induced cirrhosis, in Mdr2(Abcb4)(-/-) mice and in human end-stage liver disease. alphavbeta6 protein was absent in normal livers and expressed de novo on (activated) bile duct epithelia and transitional hepatocytes. A single dose of the alphavbeta6 inhibitor injected into Mdr2(Abcb4)(-/-) mice significantly induced profibrolytic matrix metalloproteinases (MMP)-8 and -9 after 3 h, with a corresponding increase in extracellular matrix-degrading activities. In parallel profibrogenic transcripts (procollagen alpha1(I), TGFbeta2, and MMP-2) showed a trend of downregulation. CONCLUSIONS: (1) Integrin alphavbeta6 is induced de novo in rodent and human liver fibrosis, where it is expressed on activated bile duct epithelia and (transitional) hepatocytes during fibrosis progression. (2) In vivo a single dose of a small molecule alphavbeta6 inhibitor induced antifibrogenic and profibrolytic genes and activities, suggesting alphavbeta6 is a unique target for treatment of liver fibrosis.

Integrative analysis of RUNX1 downstream pathways and target genes

BACKGROUND: The RUNX1 transcription factor gene is frequently mutated in sporadic myeloid and lymphoid leukemia through translocation, point mutation or amplification. It is also responsible for a familial platelet disorder with predisposition to acute myeloid leukemia (FPD-AML). The disruption of the largely unknown biological pathways controlled by RUNX1 is likely to be responsible for the development of leukemia. We have used multiple microarray platforms and bioinformatic techniques to help identify these biological pathways to aid in the understanding of why RUNX1 mutations lead to leukemia. RESULTS: Here we report genes regulated either directly or indirectly by RUNX1 based on the study of gene expression profiles generated from 3 different human and mouse platforms. The platforms used were global gene expression profiling of: 1) cell lines with RUNX1 mutations from FPD-AML patients, 2) over-expression of RUNX1 and CBFbeta, and 3) Runx1 knockout mouse embryos using either cDNA or Affymetrix microarrays. We observe that our datasets (lists of differentially expressed genes) significantly correlate with published microarray data from sporadic AML patients with mutations in either RUNX1 or its cofactor, CBFbeta. A number of biological processes were identified among the differentially expressed genes and functional assays suggest that heterozygous RUNX1 point mutations in patients with FPD-AML impair cell proliferation, microtubule dynamics and possibly genetic stability. In addition, analysis of the regulatory regions of the differentially expressed genes has for the first time systematically identified numerous potential novel RUNX1 target genes. CONCLUSION: This work is the first large-scale study attempting to identify the genetic networks regulated by RUNX1, a master regulator in the development of the hematopoietic system and leukemia. The biological pathways and target genes controlled by RUNX1 will have considerable importance in disease progression in both familial and sporadic leukemia as well as therapeutic implications.

Highly efficient in vivo agonist-induced internalization of sst2 receptors in somatostatin target tissues

The successful peptide receptor imaging of tumors, as exemplified for somatostatin receptors, is based on the overexpression of peptide receptors in selected tumors and the high-affinity binding to these tumors of agonist radioligands that are subsequently internalized into the tumor cells in which they accumulate. Although in vitro studies have shown ample evidence that the ligand-receptor complex is internalized, in vivo evidence of agonist-induced internalization of peptide receptors, such as somatostatin receptors, is missing. METHODS: Rats subcutaneously transplanted with the somatostatin receptor subtype 2 (sst(2))-expressing AR42J tumor cells were treated with intravenous injections of various doses of the sst(2) agonist [Tyr(3), Thr(8)]-octreotide (TATE) or of the sst(2) antagonist 1,4,7,10-tetraazacyclododecane-N,N',N'',N''',-tetraacetic acid (DOTA)-Bass and were sacrificed at various times ranging from 2.5 min to 24 h after injection. The tumors and pancreas were then removed from each animal. All tissue samples were processed for sst(2) immunohistochemistry using sst(2)-specific antibodies. RESULTS: Compared with the sst(2) receptors in untreated animals, which localized at the plasma membrane in pancreatic and AR42J tumor cells, the sst(2) receptors in treated animals are detected intracellularly after an intravenous injection of the agonist TATE. Internalization is fast, as the receptors are already internalizing 2.5 min after TATE injection. The process is extremely efficient, as most of the cell surface receptors internalize into the cell and are found in endosomelike structures after TATE injection. The internalization is most likely reversible, because 24 h after injection the receptors are again found at the cell surface. The process is also agonist-dependent, because internalization is seen with high-affinity sst(2) agonists but not with high-affinity sst(2) antagonists. The same internalization properties are seen in pancreatic and AR42J tumor cells. They can further be confirmed in vitro in human embryonic kidney-sst(2) cells, with an immunofluorescence microscopy-based sst(2) internalization assay. CONCLUSION: These animal data strongly indicate that the process of in vivo sst(2) internalization after agonist stimulation is fast, extremely efficient, and fully functional under in vivo conditions in neoplastic and physiologic sst(2) target tissues. This molecular process is, therefore, likely to be responsible for the high and long-lasting uptake of sst(2) radioligands seen in vivo in sst(2)-expressing tumors.

Cathepsin W expressed exclusively in CD8+ T cells and NK cells, is secreted during target cell killing but is not essential for cytotoxicity in human CTLs

OBJECTIVE: Cathepsin W (CatW, lymphopain) is a putative cysteine protease with restricted expression to natural killer (NK) cells and CD8(+) T cells and so far unknown function and properties. Here, we characterize in detail, the regulation of human CatW during T-cell development in response to different stimuli and its functional involvement in cytotoxic lymphocyte effector function. MATERIALS AND METHODS: Western blots and real time polymerase chain reaction of sorted, unstimulated, and stimulated cell subsets (thymocytes, T cells, NK cells) and their culture supernatants were used to study regulation and expression of CatW. Primary CD8(+) T cells and short-term T-cell lines were transfected with small interfering RNA to study the involvement of CatW in effector function such as target cell killing and interferon-gamma production. RESULTS: Levels of CatW expression correlate closely with cytotoxic capacity both during development and in response to factors influencing cytotoxicity. Furthermore, CatW is secreted during specific target cell killing. However, knockdown of CatW expression by small interfering RNA neither influences target cell killing nor interferon-gamma production. CONCLUSION: Despite being expressed in the effector subset of CD8(+) and NK cells and of being released during target cell killing, our functional inhibition studies exclude an essential role of CatW in the process of cytotoxicity.

Patterns of recurrence of early breast cancer according to estrogen receptor status: a therapeutic target for a quarter of a century

The current therapeutic strategy in breast cancer is to identify a target, such as estrogen receptor (ER) status, for tailoring treatments. We investigated the patterns of recurrence with respect to ER status for patients treated in two randomized trials with 25 years' median follow-up. In the ER-negative subpopulations most breast cancer events occurred within the first 5-7 years after randomization, while in the ER-positive subpopulations breast cancer events were spread through 10 years. In the ER-positive subpopulation, 1 year endocrine treatment alone significantly prolonged disease-free survival (DFS) with no additional benefit observed by adding 1 year of chemotherapy. In the small ER-negative subpopulation chemo-endocrine therapy had a significantly better DFS than endocrine alone or no treatment. Despite small numbers of patients, "old-fashioned" treatments, and competing causes of treatment failure, the value of ER status as a target for response to adjuvant treatment is evident through prolonged follow-up.