Magnetic Tracking and Electrocardiography-Guided Tip Confirmation System Versus Fluoroscopy for Placement of Peripherally Inserted Central Catheters: A Randomized, Noninferiority Comparison.

PURPOSE: To determine whether the use of a magnetic tracking and electrocardiography-guided catheter tip confirmation system (TCS) is safe and noninferior to fluoroscopy concerning positioning accuracy of a peripheral inserted central catheter (PICC). METHODS: In this prospective, randomized, single-center study, adult patients scheduled for PICC insertion were assigned 1:1 either to TCS or fluoroscopy. The primary objective was a noninferiority comparison of correct PICC tip position confirmed by X-ray obtained immediately after catheter insertion. Time needed for PICC insertion and insertion-related complications up to 14 days after the procedure were secondary outcomes to be assessed for superiority. RESULTS: A total of 210 patients (62.3 ± 14.4 years, 63.8% male) were included at a single German center between June 2016 and October 2017. Correct PICC tip position was achieved in 84 of 103 TCS (82.4%) and 103 of 104 fluoroscopy patients (99.0%). One-sided 95% lower confidence limit on the difference between proportions was -23.1%. Thus, noninferiority of TCS was not established (p > 0.99). Insertion of PICC took longer with TCS compared to fluoroscopy (8.4 ± 3.7 min vs. 5.0 ± 2.7 min, p < 0.001). Incidence of complications within a mean follow-up of 5.0 ± 2.3 days did not differ significantly between groups. CONCLUSION: Noninferiority of TCS to fluoroscopy in the incidence of correct PICC tip position was not reached. Ancillary benefit of TCS over fluoroscopy including less radiation exposure and lower resource requirements may nonetheless justify the use of TCS. The study is registered with Clinical.Trials.gov (Identifier: NCT02929368).

Restoration of spatial working memory by genetic rescue of GluR-A-deficient mice.

Gene-targeted mice lacking the AMPA receptor subunit GluR-A (also called GluR1 encoded by the gene Gria1,) have deficits in hippocampal CA3-CA1 long-term potentiation (LTP) and have profoundly impaired hippocampus-dependent spatial working memory (SWM) tasks, although their spatial reference memory remains normal. Here we show that forebrain-localized expression of GFP-tagged GluR-A subunits in GluR-A-deficient mice rescues SWM, paralleling its rescue of CA3-CA1 LTP. This provides powerful new evidence linking hippocampal GluR-A-dependent synaptic plasticity to rapid, flexible memory processing.

Molecular imaging of carcinogenesis with immuno-targeted nanoparticles.

Molecular characterization of cancer could have important clinical benefits such as earlier cancer detection based on molecular characterization, the ability to predict the risk of cancer progression, real time margin detection, the ability to rationally select molecular therapy and to monitor response to the therapy. We present a new class of molecular specific contrast agents for optical imaging of carcinogenesis in vivo - gold nanoparticles conjugated with monoclonal antibodies specific for cancer biomarkers.

Dominance of the lurcher mutation in heteromeric kainate and AMPA receptor channels.

Homomeric glutamate receptor (GluR) channels become spontaneously active when the last alanine residue within the invariant SYTANLAAF-motif in the third membrane segment is substituted by threonine. The same mutation in the orphan GluRdelta2 channel is responsible for neurodegeneration in "Lurcher" (Lc) mice. Since most native GluRs are composed of different subunits, we investigated the effect of an Lc-mutated subunit in heteromeric kainate and AMPA receptors expressed in HEK293 cells. Kainate receptor KA2 subunits, either wild type or carrying the Lc mutation (KA2(Lc)), are retained inside the cell but are surface-expressed when assembled with GluR6 subunits. Importantly, KA2(Lc) dominates the gating of KA2(Lc)/GluR6(WT) channels, as revealed by spontaneous activation and by slowed desensitization and deactivation kinetics of ligand-activated whole-cell currents. Moreover, the AMPA receptor subunit GluR-B(Lc)(Q) which forms spontaneously active homomeric channels with rectifying current-voltage relationships, dominates the gating of heteromeric GluR-B(Lc)(Q)/GluR-A(R) channels. The spontaneous currents of these heteromeric AMPAR channels show linear current-voltage relationships, and the ligand-activated whole-cell currents display slower deactivation and desensitization kinetics than the respective wild-type channels. For heteromeric Lc-mutated kainate and AMPA receptors, the effects on kinetics were reduced relative to the homomeric Lc-mutated forms. Thus, an Lc-mutated subunit can potentially influence heteromeric channel function in vivo, and the severity of the phenotype will critically depend on the levels of homomeric GluR(Lc) and heteromeric GluR(Lc)/GluR(WT) channels.

Comparative analysis of IFN-gamma B7.1 and antisense TGF-beta gene transfer on the tumorigenicity of a poorly immunogenic metastatic mammary carcinoma.

Cancer progression is attributed in part to immune evasion strategies that include lack of co-stimulation, down-regulation of cell surface MHC molecules, and secretion of immunosuppressive factors, such as transforming growth factor-beta (TGF-beta). Gene therapy has been employed to counter these mechanisms of immune evasion by transference of B7.1, IFN-gamma or antisense TGF-beta genes into tumor cells, resulting in cell surface expression of B7.1, upregulation of MHC class I and class II molecules, or elimination of tumor-derived TGF-beta, respectively. Although each of these transgenes has been shown to alter tumorigenicity in murine models, a direct comparison of their efficacy has not been performed. In this study, we have employed a very aggressive, poorly immunogenic and highly metastatic mammary model, 4T1, to compare the efficacy of B7.1, IFN-gamma and antisense TGF-beta gene transfer in stimulating an anti-tumor response. We demonstrate that both IFN-gamma and antisense TGF-beta gene expression significantly reduced the tumorigenicity of these cells compared to mock transduced cells, with IFN-gamma having a greater effect. In contrast, B7.1 gene transfer did not affect the tumorigenicity of 4T1 cells. The anti-tumor response directed against antisense TGF-beta-expressing 4T1 tumors was mediated by CD4+ and CD8+ T cells. However, CD8+ T cells, and not CD4+ T cells, appeared to mediate the anti-tumor response against IFN-gamma-expressing tumors. Treatment of tumor-bearing animals with IFN-gamma or antisense TGF-beta gene-modified tumor cell vaccines reduced the number of clonogenic metastases to the lungs and liver compared to treatment with mock-transduced cells. Finally, in a residual disease model in which the primary tumor was excised and mice were vaccinated with irradiated tumor cells, treatment of mice with vaccinations consisting of 4T1 cells expressing both antisense TGF-beta and IFN-gamma genes was the most effective in prolonging survival.

Macrophage effector mechanisms in melanoma in an experimental study.

BACKGROUND: The tumor-bearing state is known to induce immune dysfunction that contributes to increased infectious complications and tumor progression. However, the mechanisms underlying this immunosuppression remain unclear. HYPOTHESIS: Macrophage (MO) dysfunction may play a role in tumor-induced immunosuppression. DESIGN AND MAIN OUTCOME MEASURES: Using a murine model, this study investigated the effects of melanoma growth on peritoneal macrophage effector molecule and prostaglandin production, MO-mediated cytotoxicity, and candidacidal mechanisms. Female C57BL/6 mice were inoculated with 106 B16 melanoma cells or a salt solution subcutaneously. Mice were euthanized 3 weeks later and peritoneal MOs were harvested and assayed for nitric oxide, superoxide anion, tumor necrosis factor alpha, and prostaglandin E(2)production. Macrophage-mediated cytotoxicity against B16 melanoma targets and MO candidacidal mechanisms were also measured. RESULTS: Macrophage production of nitric oxide, superoxide anion, and tumor necrosis factor alpha were significantly decreased, while prostaglandin E(2)production was increased in MOs from melanoma-bearing mice. Concomitantly, MO-mediated cytotoxicity and candidacidal mechanisms were significantly impaired. CONCLUSIONS: Melanoma growth leads to decreased MO effector molecule production, increased prostaglandin E(2)production, and impaired MO cytotoxic and candidacidal mechanisms. These results may help explain the observed increased infectious complications in the tumor-bearing host. Strategies aimed at restoring MO function may have therapeutic potential.

Suppression of macrophage nitric oxide production by melanoma: mediation by a melanoma-derived product.

The tumour-bearing state is known to induce immune dysfunction that contributes to increased infectious complications and tumour progression. However, the mechanisms underlying this immunosuppression remain unclear. This study investigated in a murine model the effects of melanoma growth on nitric oxide (NO) production by peritoneal macrophages in vivo and in vitro. B16 and K1735 melanoma cells were inoculated subcutaneously into C57BL/6 and C3H/HeN mice, respectively. Stimulated NO production by elicited peritoneal macrophages was examined in control and melanoma- bearing mice. An in vitro system was established to assess the effects of co-culturing melanoma cells (B16 and K1735) or melanoma-conditioned medium with normal peritoneal macrophages on subsequent NO production. NO production was significantly suppressed in macrophages from melanoma-bearing mice. Co-culture of normal macrophages with melanoma cells in a transwell system or with melanoma-conditioned media in vitro reproduced the defects observed in vivo without affecting macrophage viability, pointing to a melanoma-derived product as the basis for the observed suppression of NO production. This inhibition required RNA and protein synthesis and was dose and time dependent. Using inhibition profiles and neutralizing antibodies, it was demonstrated that this melanoma inhibitory activity was distinct from known NO inhibitors. Preliminary characterization attributed this activity to a melanoma-secreted protein moiety.

Conditional restoration of hippocampal synaptic potentiation in Glur-A-deficient mice.

Plasticity of mature hippocampal CA1 synapses is dependent on l-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors containing the glutamate receptor A (GluR-A) subunit. In GluR-A-deficient mice, plasticity could be restored by controlled expression of green fluorescent protein (GFP)-tagged GluR-A, which contributes to channel formation and displayed the developmental redistribution of AMPA receptors in CA1 pyramidal neurons. Long-term potentiation (LTP) induced by pairing or tetanic stimulation was rescued in adult GluR-A(-/-) mice when (GFP)GluR-A expression was constitutive or induced in already fully developed pyramidal cells. This shows that GluR-A-independent forms of synaptic plasticity can mediate the establishment of mature hippocampal circuits that are prebuilt to express GluR-A-dependent LTP.

Invasion and metastasis of a mammary tumor involves TGF-beta signaling.

Several studies have correlated escape from TGF-beta-mediated cell cycle arrest with the tumorigenic phenotype. Most often, this escape from growth control has been linked to dysfunctional TGF-beta receptors or defects in the TGF-beta-mediated SMAD signaling pathway. In this report, we found that highly metastatic 4T1 mammary carcinoma cells express functional TGF-beta receptors capable of initiating SMAD-mediated transcription, yet are not growth inhibited by TGF-beta1. We further observed that TGF-beta directly contributes to the metastatic behavior of this cell line. Exposure to TGF-beta caused 4T1 cells to undergo morphological changes associated with the metastatic phenotype and invade more readily through collagen coated matrices. Furthermore, expression of a dominant negative truncated type II receptor diminished TGF-beta signaling and significantly restricted the ability of 4T1 cells to establish distant metastases. Our results suggest that regardless of 4T1 resistance to TGF-beta-mediated growth inhibition, TGF-beta signaling is required for tumor invasion and metastases formation.

Trauma-induced alterations in macrophage function.

BACKGROUND: The juxtaposition of immune suppression and a hyperactive inflammatory response after injury represents a paradox in immune function. The aim of this study was to evaluate the delayed macrophage hypersecretion of inflammatory mediators in relation to functional macrophage defects. METHODS: BALB/c mice were randomized to control or trauma (femur fracture plus 40% blood volume hemorrhage) groups. One and 7 days after injury, splenic macrophages were isolated and assayed for antigen presentation and the production of inflammatory mediators tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-6, prostaglandin E2, H2O2, and nitric oxide. RESULTS: One day after injury, there were significantly diminished macrophage antigen presentation and decreased mean production of TNF-alpha, IL-6, and H2O2. In contrast, 7 days after injury, splenic macrophages produced significantly increased mean amounts of TNF-alpha, IL-6, prostaglandin E2, H2O2, and nitric oxide, with a persistent functional defect in antigen presentation. CONCLUSIONS: This phasic response to trauma suggests a persistent state of macrophage dysregulation that may help explain the paradox of immune suppression, manifested by functional defects predisposing patients to increased infections, in the setting of inflammatory mediator hypersecretion, predisposing patients to the systemic inflammatory response syndrome/multiple organ dysfunction syndrome.

Candida infection following severe trauma exacerbates Th2 cytokines and increases mortality.

Following trauma, there is an increase of Th2 cytokines (IL-4, IL-6, and IL-10) and a decrease in Th1 cytokines (IFN-gamma and IL-2) that may account for impaired cellular immunity. However, the functional significance of a dominant Th2 pattern to the host remains unclear. The aim of this study was to evaluate whether Candida albicans (CA) sepsis in the setting of a Th2 response to trauma leads to increased mortality and to examine the mediators involved. Female BALB/c mice were randomized (12 per group) to receive no injury (C); trauma, consisting of a combined femur fracture and 40% total blood loss (T); no injury plus CA infection (C+CA); and CA infection 1 week following trauma (T+CA). Survival was then followed for 3 weeks. In a separate study, mice were treated as above (5 per group) and sacrificed. Harvested splenocytes were evaluated for concanavalin A-stimulated cytokine production and liver and kidney homogenates were plated to evaluate CA growth per organ and examined histologically. Candida infection at 1 week following trauma resulted in significantly increased mortality compared to infected controls. Furthermore, the Th2 dominant cytokine pattern was significantly augmented in the presence of CA infection in both C+CA and T+CA groups. Additional analysis showed significant growth of CA in liver and kidney homogenates from T+CA compared to C+CA mice. These results suggest that injured and infected mice demonstrate augmentation of Th2 dominant responses above that of injury or infection alone, as well as a decreased ability to clear Candida which may partially explain the increase in mortality observed. Therapies designed to neutralize Th2 cytokines or augment Th1 cytokines may prove beneficial in the setting of sepsis following trauma.

Dominance of T-helper 2-type cytokines after severe injury.

OBJECTIVE: To determine whether severe injury leads to a dominance of splenocyte-produced T-helper (Th) 2-type cytokines, partly explaining the observed defects in cellular immune responses in the posttraumatic state. DESIGN: Female BALB/c mice (n = 6 per group) were randomized to receive anesthesia alone (control) or a combined femur fracture and a hemorrhage of 40% of total blood volume (trauma). On days 1 and 7 after injury, mice were killed and spleens were harvested. Splenocytes were stimulated in vitro with 2.5 micrograms of concanavalin A per milliliter. After 72 hours of incubation, splenocyte proliferation was determined by means of tritiated thymidine uptake. Production of interferon-gamma and interleukins (IL) -2, -4, -5, -6, and -10 from supernatants harvested after 24 or 72 hours of incubation was quantified by enzyme-linked immunosorbent assay. SETTING: Surgical immunology research laboratory of a medical college. MAIN OUTCOME MEASURES: Mouse spleen weight, splenocyte number, and proliferation in addition to cytokine production (interferon-gamma, IL-2, IL-4, IL-5, IL-6, and IL-10). RESULTS: Splenocyte proliferative capacity was unaffected at day 1 after injury but was significantly suppressed (P < .05) by day 7 after injury. Similarly, there were no changes in splenocyte cytokine production in a comparison of control and injured mice at day 1. At day 7, however, there was nearly a 90% decrease in the Th1-type cytokines (interferon-gamma and IL-2; P < or = .002) and at least a 30% increase in the Th2-type cytokines IL-4, IL-5, IL-6, and IL-10 (P = .06 for IL-6 and P < or = .03 for IL-4, IL-5, and IL-10). CONCLUSIONS: These data indicate that a shift to a Th2-type splenocyte cytokine response occurs late, at 7 days after injury. Modulation of Th cell cytokine responses may partially explain defects observed in cellular immune responses in postinjury states. Therapies that augment Th1-type cytokine production and/or neutralize Th2-type cytokines may prove beneficial.

Effect of fasting on hepatocytes cold stored in University of Wisconsin solution for 24 hours.

Although there have been improvements in liver preservation, liver dysfunction still remains a serious consequence of liver transplantation. This may be related to cold ischemic injury since the incidence of dysfunction increases with longer preservation times. However, even some livers preserved for short periods of time (less than 15 hr) develop liver dysfunction. One possible cause may be the lack of adequate nutritional support, and the donor may be exposed to prolonged periods of hyponutrition. In this study, we have compared the effects of fasting on functions of hepatocytes isolated from the rat. Hepatocytes were cold stored in University of Wisconsin solution for 24 hr and analyzed at the end of preservation as well as at the end of rewarming in Krebs-Henseleit buffer for 120 min. The glycogen content of fed cells was 1.57 mumol/mg protein and this was reduced by 95% in cells from fasted rats. After cold storage and rewarming, hepatocytes from fasted rats lost 84.2 +/- 2.5% of the total cellular lactate dehydrogenase versus only 32.7 +/- 3.8% (P < 0.001) in cells from fed rats. Also, ATP and reduced glutathione content of fasted cells were significantly reduced, free fatty acids were higher (P = 0.0154), and protein synthesis was reduced to 41% of controls (versus only 88% in fed cells), although there were no differences in phospholipid content. When hepatocytes from fasted rats were rewarmed in Krebs-Henseleit buffer containing fructose (10 mM), lactate dehydrogenase release was reduced from 80% to 34.4 +/- 0.2% and ATP content was significantly higher with fructose than without. Hepatocytes from fasted rats, therefore, are more sensitive to cold ischemic injury than cells from fed rats. The increased sensitivity appears related to the lack of glycogen as a source of substrates for metabolism during rewarming. This is supported by the fact that addition of fructose, which is metabolized readily by hepatocytes through glycolysis, suppressed rewarming injury to cells from fasted rats. The nutritional status of the donor, therefore, may play a pivotal role in the results of liver preservation and transplantation. Effective donor nutritional management may reduce the incidence of liver dysfunction after transplantation.

Glycine protects hepatocytes from injury caused by anoxia, cold ischemia and mitochondrial inhibitors, but not injury caused by calcium ionophores or oxidative stress.

Isolated hepatocytes, suspended in an organ preservation solution, can be preserved at 4 degrees C for up to 6 days. After preservation, normothermic-normoxic incubation causes loss of hepatocyte viability. The addition of 3 mmol/L glycine to the rewarming medium prevents the loss of viability. In this study we investigated the cytoprotective effects of glycine under many conditions known to cause hepatocellular injury to understand the mechanism of cold-induced injury in the liver. Hepatocytes were suspended in modified Krebs-Henseleit buffer with or without 3 mmol/L glycine and exposed to agents or conditions known to induce cell death. Hepatocyte viability was assessed by measuring the percentage of lactate dehydrogenase leakage from the cells and the concentration of ATP during incubation at 37 degrees C under room air for up to 90 min. Mitochondrial inhibitors (potassium cyanide and carbonyl cyanide m-chlorophenylhydrazone); calcium ionophores (ionomycin and A23187); an oxidizing agent, tert-butyl hydroperoxide; and anoxia were all used to cause cell injury. Hepatocytes were also isolated from fasted rats and hypothermically preserved as another model of cell death. Other amino acids were also tested in the hypothermic preservation model to study the specificity of the amino acid requirement for prevention of lactate dehydrogenase leakage. Of the amino acids tested, only alanine (10 mmol/L) and the combination of alanine (3 mmol/L) and serine (3 mmol/L) were as effective as glycine in preventing lactate dehydrogenase release in the hypothermic preservation model.(ABSTRACT TRUNCATED AT 250 WORDS)