Intracellular survival of virulence and low-virulence strains of Vibrio parahaemolyticus in Epinephelus awoara macrophages and peripheral leukocytes.

In this study, we examined the virulence factors and pathogenesis of Vibrio parahaemolyticus in Epinephelus awoara. The chemotactic motility of V. parahaemolyticus for phagocytosis and intracellular survival in fish macrophages was determined using virulence strains and low-virulence strains of V. parahaemolyticus. We found that the intracellular mean number of virulence strains of V. parahaemolyticus ranged from 0-180 min after co-incubation with macrophages and peripheral leukocytes, was relatively low, and decreased steadily over the observation period. Low-virulence strains of V. parahaemolyticus were unable to survive in peripheral leukocytes and macrophages. Cell viability in response to V. parahaemolyticus was assessed using the MTT assay. Low-virulence V. parahaemolyticus strains exhibited lower cytotoxicity compared to virulent strains. The average percent of live macrophages and peripheral leukocytes infected by V. parahaemolyticus ranged from 13.50-79.20%. These results indicate that V. parahaemolyticus in E. awoara is a facultative intracellular bacterium that may be involved in virulence.

Interleukin-32 enhances cytotoxic effect of natural killer cells to cancer cells via activation of death receptor 3.

Studies have demonstrated that the anti-tumour effect of natural killer (NK) cells is successful for patients with several cancers. Although interleukin-32 (IL-32) is endogenously expressed in NK cells, cytolytic function of NK cells against cancer cells has not been fully demonstrated. In the present study, we found that the growth of cancer cells was suppressed when colon cancer cells or prostate cancer cells were co-cultured with NK-92 cells, an NK cell line. We also found that the expression of tumour necrosis factor receptor 2 and death receptor 3 (DR3) was increased in PC3 cells, and the expression of FAS and DR3 was increased in SW620 cells by co-culture with NK-92 cells. However, cancer cell growth inhibition and IL-32 expression were abolished when cancer cells were co-cultured with NK cells transfected with small interfering (si) RNA of IL-32. DR3 expression was also diminished by co-culture with IL-32-specific siRNA-transfected NK-92 cells. Expression of APO3L, a ligand of DR3, was elevated in NK cells that were co-cultured with cancer cells. It was also found that expression of apoptosis-related proteins such as cleaved caspase-3 and bax was increased in cancer cells co-cultured with NK-92 cells, but their expression was abolished by co-culture with IL-32 siRNA-transfected NK-92 cells. Moreover, knockdown of DR3 in co-culture of NK-92 cells with cancer cells by siRNA or antibodies of DR3 and APO3L reversed the growth inhibitory effect of NK-92 cells. In conclusion, our study showed that IL-32 enhanced the cytotoxic effect of NK-92 cells on the cancer cells through activation of DR3 and caspase-3.

Comparison of nerve growth factor receptor binding models using heterodimeric muteins.

Nerve growth factor (NGF) is a homodimer that binds to two distinct receptor types, TrkA and p75, to support survival and differentiation of neurons. The high-affinity binding on the cell surface is believed to involve a heteroreceptor complex, but its exact nature is unclear. We developed a heterodimer (heteromutein) of two NGF muteins that can bind p75 and TrkA on opposite sides of the heterodimer, but not two TrkA receptors. Previously described muteins are Δ9/13 that is TrkA negative and 7-84-103 that is signal selective through TrkA. The heteromutein (Htm1) was used to study the heteroreceptor complex formation and function, in the putative absence of NGF-induced TrkA dimerization. Cellular binding assays indicated that Htm1 does not bind TrkA as efficiently as wild-type (wt) NGF but has better affinity than either homodimeric mutein. Htm1, 7-84-103, and Δ9/13 were each able to compete for cold-temperature, cold-chase stable binding on PC12 cells, indicating that binding to p75 was required for a portion of this high-affinity binding. Survival, neurite outgrowth, and MAPK signaling in PC12 cells also showed a reduced response for Htm1, compared with wtNGF, but was better than the parent muteins in the order wtNGF > Htm1 > 7-84-103 >> Δ9/13. Htm1 and 7-84-103 demonstrated similar levels of survival on cells expressing only TrkA. In the longstanding debate on the NGF receptor binding mechanism, our data support the ligand passing of NGF from p75 to TrkA involving a transient heteroreceptor complex of p75-NGF-TrkA.

Ligand-dependent TrkA activity in brain differentially affects spatial learning and long-term memory.

In the central nervous system, the nerve growth factor (NGF) receptor TrkA is expressed primarily in cholinergic neurons that are implicated in spatial learning and memory, whereas the NGF receptor p75(NTR) is expressed in many neuronal populations and glia. We asked whether selective TrkA activation may have a different impact on learning, short-term memory, and long-term memory. We also asked whether TrkA activation might affect cognition differently in wild-type mice versus mice with cognitive deficits due to transgenic overexpression of mutant amyloid-precursor protein (APP mice). Mice were treated with wild-type NGF (a ligand of TrkA and p75(NTR)) or with selective pharmacological agonists of TrkA that do not bind to p75(NTR). In APP mice, the selective TrkA agonists significantly improved learning and short-term memory. These improvements are associated with a reduction of soluble Aß levels in the cortex and AKT activation in the cortex and hippocampus. However, this improved phenotype did not translate into improved long-term memory. In normal wild-type mice, none of the treatments affected learning or short-term memory, but a TrkA-selective agonist caused persistent deficits in long-term memory. The deficit in wild-type mice was associated temporally, in the hippocampus, with increased AKT activity, increased brain-derived neurotrophic factor precursor, increased neurotrophin receptor homolog-2 (p75-related protein), and long-term depression. Together, these data indicate that selective TrkA activation affects cognition but does so differently in impaired APP mice versus normal wild-type mice. Understanding mechanisms that govern learning and memory is important for better treatment of cognitive disorders.

Chronic and acute models of retinal neurodegeneration TrkA activity are neuroprotective whereas p75NTR activity is neurotoxic through a paracrine mechanism.

In normal adult retinas, NGF receptor TrkA is expressed in retinal ganglion cells (RGC), whereas glia express p75(NTR). During retinal injury, endogenous NGF, TrkA, and p75(NTR) are up-regulated. Paradoxically, neither endogenous NGF nor exogenous administration of wild type NGF can protect degenerating RGCs, even when administered at high frequency. Here we elucidate the relative contribution of NGF and each of its receptors to RGC degeneration in vivo. During retinal degeneration due to glaucoma or optic nerve transection, treatment with a mutant NGF that only activates TrkA, or with a biological response modifier that prevents endogenous NGF and pro-NGF from binding to p75(NTR) affords significant neuroprotection. Treatment of normal eyes with an NGF mutant-selective p75(NTR) agonist causes progressive RGC death, and in injured eyes it accelerates RGC death. The mechanism of p75(NTR) action during retinal degeneration due to glaucoma is paracrine, by increasing production of neurotoxic proteins TNF-α and α(2)-macroglobulin. Antagonists of p75(NTR) inhibit TNF-α and α(2)-macroglobulin up-regulation during disease, and afford neuroprotection. These data reveal a balance of neuroprotective and neurotoxic mechanisms in normal and diseased retinas, and validate each neurotrophin receptor as a pharmacological target for neuroprotection.

Cranberry extract-based formulations for preventing bacterial biofilms.

Generating formulations for the delivery of a mixture of natural compounds extracted from natural sources is a challenge because of unknown active and inactive ingredients and possible interactions between them. As one example, natural cranberry extracts have been proposed for the prevention of biofilm formation on dental pellicle or teeth. However, such extracts may contain phenolic acids, flavonol glycosides along with other constituents like coumaroyl iridoid glycosides, flavonoids, alpha-linolenic acid, n-6 (or n-3) fatty acids, and crude fiber. Due to the presence of a variety of compounds, determining which molecules (and how many molecules) are essential for preventing biofilm growth is nontrivial to ascertain. Therefore, a formulation that could contain natural, unrefined, cranberry extract (with all its constituent compounds) at high loading would be ideal. Accordingly, we have generated several candidate formulations including poly(lactic-co-glycolic) acid (PLGA)-based microencapsulation of cranberry extract (CE15) as well as formulations including stearic acid along with polyvinylpyrrolidone (PVP) or Ethyl lauroyl arginate (LAE) complexed with cranberry extracts (CE15). We found that stearic acid in combination with PVP or LAE as excipients led to higher loading of the active and inactive compounds in CE15 as compared with a PLGA microencapsulation and also sustained release of CE15 in a tunable manner. Using this method, we have been able to generate two successful formulations (one preventative based, one treatment based) that effectively inhibit biofilm growth when incubated with saliva. In addition to cranberry extract, this technique could also be a promising candidate for other natural extracts to form controlled release systems.Graphical abstract.

Identification of critical residues within the conserved and specificity patches of nerve growth factor leading to survival or differentiation.

Afflicted neurons in Alzheimer disease have been shown to display an imbalance in the expression of TrkA and p75(NTR) at the cell surface, and administration of nerve growth factor (NGF) has been considered and attempted for treatment. However, wild-type NGF causes extensive elaboration of neurites while providing survival support. This study was aimed at developing recombinant NGF muteins that did not support neuritogenesis while maintaining the survival response. Critical residues were identified at the ligand-receptor interface by point mutagenesis that played a greater importance in neuritogenesis versus survival. By combining point mutations, two survival-selective recombinant NGF muteins, i.e./7-84-103 and KKE/7-84-103, were generated. Both muteins reduced neuritogenesis in PC12 (TrkA(+)/p75(NTR+)) cells by >90%, while concurrently retaining near wild-type survival activity in MG139 (TrkA(+) only) and PCNA fibroblast (p75(NTR+)-only) cells. Additionally, survival in both naive and terminally differentiated PC12 cells was shown to be intermediate between NGF and negative controls. Dose-response curves with 7-84-103 showed that the differentiation curve was shifted by about 100-fold, whereas the EC(50) for survival was only increased by 3.3-fold. Surface plasmon resonance analysis revealed a 200-fold decrease in binding of 7-84-103 to TrkA. The retention of cell survival was attributed to maintenance of signaling through the Akt survival pathway with reduced MAPK signaling for differentiation. The effect of key mutations along the NGF receptor interface are transmitted inside the cell to enable the generation of survival-selective recombinant NGF muteins that may represent novel pharmacologic lead agents for the amelioration of Alzheimer disease.

Organoarsenicals in Seafood: Occurrence, Dietary Exposure, Toxicity, and Risk Assessment Considerations - A Review.

Diet, especially seafood, is the main source of arsenic exposure for humans. The total arsenic content of a diet offers inadequate information for assessment of the toxicological consequences of arsenic intake, which has impeded progress in the establishment of regulatory limits for arsenic in food. Toxicity assessments are mainly based on inorganic arsenic, a well-characterized carcinogen, and arsenobetaine, the main organoarsenical in seafood. Scarcity of toxicity data for organoarsenicals, and the predominance of arsenobetaine as an organic arsenic species in seafood, has led to the assumption of their nontoxicity. Recent toxicokinetic studies show that some organoarsenicals are bioaccessible and cytotoxic with demonstrated toxicities like that of pernicious trivalent inorganic arsenic, underpinning the need for speciation analysis. The need to investigate and compare the bioavailability, metabolic transformation, and elimination from the body of organoarsenicals to the well-established physiological consequences of inorganic arsenic and arsenobetaine exposure is apparent. This review provides an overview of the occurrence and assessment of human exposure to arsenic toxicity associated with the consumption of seafood.

Absorbing phase transition in a conserved lattice gas model in one dimension.

We investigated the nonequilibrium phase transition of the conserved lattice gas model in one dimension using two update methods: i.e., the sequential update and the parallel update. We measured the critical indices of theta, beta, nu(parallel), and nu(perpendicular), and found that, for a parallel update, the exponents were delicately influenced by the hopping rule of active particles. When the hopping rule was designed to be symmetric, the results were found to be consistent with those of the sequential update. The exponents we obtained were precisely the same as the corresponding results of a recently presented lattice model of two species of particles with a conserved field in one dimension, in contrast with the authors' claim. We also found that one of the scaling relations known for absorbing phase transition is violated.

Universality split in absorbing phase transition with conserved field on fractal lattices.

The universality split in absorbing phase transition between the conserved lattice gas (CLG) model and the conserved threshold transfer process (CTTP) is investigated on a checkerboard fractal and on a Sierpinski gasket. The critical exponents theta, beta, nu||, and z, which are associated with, respectively, the density of active particles in time, the order parameter, the temporal correlation length, and the dynamics of active particles, are elaborately measured for two models on selected fractal lattices. The exponents for the CLG model are found to be distinctly different from those of the CTTP model on a checkerboard fractal, whereas the two models exhibit the same critical behavior on a Sierpinski gasket, indicating that the universality split between the two models occurs only on a checkerboard fractal. Such a universality split is attributed from the dominant hopping mechanisms caused by the intrinsic properties of the underlying fractal lattice.

Validity of scaling relations in absorbing phase transitions with a conserved field.

The two scaling relations in absorbing phase transitions, nu_||=beta/theta and z=nu_||/nu_(perpendicular), are studied for a conserved lattice gas model. The critical indices calculated elaborately from the all-sample average density of active particles appear to satisfy both relations. However, the exponent nu_(perpendicular) calculated from the surviving samples does not appear to be consistent with the value in the thermodynamic limit. This is in contrast with earlier observations [M. Rossi, Phys. Rev. Lett. 85, 1803 (2000); S. Lübeck and P. C. Heger, Phys. Rev. E. 68, 056102 (2003)], in that the former scaling relation was claimed to be violated.

Diabetes With or Without Hypertension Does Not Affect Graft Survival and All-cause Mortality After Liver Transplant: A Propensity Score Matching Analysis.

BACKGROUND: Many studies have reported the negative influence of diabetes and hypertension on morbidity and mortality in the general population. In liver transplantation (LT) recipients, prevalence of nonalcoholic fatty liver disease and metabolic syndrome is increasing. Hence, concerns over the negative influence of metabolic syndrome, including diabetes and hypertension, are growing. However, there have been few studies about the outcomes of LT recipients with diabetes with/without hypertension. We aimed to evaluate the impact of diabetes with/without hypertension on the outcomes of LT. METHODS: Between May 2010 and October 2015, 814 LT recipients (median age, 51 [46-55] years; median MELD score, 13 [9-18]), without overt cardiovascular disease were retrospectively evaluated. To rigorously adjust for clinically confounding factors, a 1:2 propensity score matching analysis was performed. Kaplan-Meier survival curves and Cox proportional hazard regression analysis were performed to examine the association between diabetes with/without hypertension and all-cause mortality or graft survival rate. RESULTS: There were 77 (9.5%) graft failures and 71 (8.7%) deaths during a median follow-up of 2.4 years. After 1:2 matching of 173 (21.3%) diabetic patients, no significant differences were evident in graft survival rate (log-rank test, P = .46; and hazard ratio [HR], 1.06; 95% confidence interval [CI], 0.55-2.06; P = .865) and all-cause mortality (log-rank test, P = .59; and HR, 1.06; 95% CI, 0.55-2.06; P = .727). Separate 1:2 matching was applied to a subgroup of 43 (5.3%) patients with diabetes and hypertension. This matching also showed no difference in graft survival rate (log-rank test, P = .45; and HR, 1.35; 95% CI, 0.43-4.27; P = .613) and all-cause mortality (log-rank test, P = .25; and HR, 1.87; 95% CI, 0.54-6.50; P = .325). CONCLUSION: Diabetes with/without hypertension does not have an impact on graft survival rate or all-cause mortality in LT recipients.

Effects of chronic statin use on 30-day major adverse cardiac and cerebrovascular events after thoracic endovascular aortic repair.

BACKGROUND: Cardiac and cerebrovascular complications are major causes of adverse outcomes following thoracic endovascular aortic repair (TEVAR). The benefits of statins have been established, but little is known about their impact on patients undergoing TEVAR. We investigated whether chronic statin use protected against early postoperative major adverse cardiac and cerebrovascular events (MACCEs) after TEVAR. METHODS: We retrospectively reviewed 211 patients who underwent TEVAR between February 2013 and March 2017 classified into two groups, those with acute aortic syndrome (AAS, N.=79) and those without (non-AAS, N.=132). Patients were subdivided according to preoperative statin therapy for ≥3 months or not. The primary endpoint was 30-day MACCE, defined as myocardial infarction, stroke, arrhythmia, cardiovascular death, or cerebrovascular death. Acute kidney injury (AKI) occurrence within 48 hours was also evaluated. Multivariate logistic regression analysis was performed to identify independent risk factors for MACCEs and AKI. RESULTS: Incidence of MACCEs (1% vs. 11%, P=0.019) was significantly lower in the statin group than in the no-statin group in non-AAS patients. Multivariate logistic regression analysis revealed statin use (odds ratio 0.85, 95% confidence interval 0.01-0.95, P=0.046) as an independent predictor for MACCE in non-AAS patients. The AKI incidence was significantly higher in the statin group than in the no-statin group in AAS patients (44% vs. 15%, P=0.018). CONCLUSIONS: In patients undergoing TEVAR, chronic statin use was associated with reduced 30-day MACCEs in non-AAS patients, but not in AAS patients. It might rather be associated with increased risk of AKI in AAS patients.

Adjacent crosstalk suppression in a colorless WDM passive optical network.

We propose and experimentally demonstrate a novel wavelength-division- multiplexed passive optical network (WDM-PON) scheme for the suppression of adjacent crosstalk arising from the wavelength misalignment in arrayed waveguide gratings (AWGs) between a central office (CO) and a remote node (RN). The adjacent crosstalk suppression is achieved by allocating two different bands to adjacent channels of the AWGs by utilizing interleavers and WDM filters. The transmission performance of the proposed scheme was measured at a 155 Mb/s data stream, and error free transmission with a power penalty less than 0.7 dB was successfully achieved in case of AWG misalignment of 0.3 nm.

Absorbing phase transition in conserved lattice gas model on fractal lattices.

We study the continuous phase transition of the conserved lattice gas (CLG) model from an active phase into an absorbing phase on two fractal lattices, i.e., on a checkerboard fractal and on a Sierpinski gasket. In the CLG model, a particle is assumed to be active if any of the neighboring sites are occupied by a particle and inactive if all neighboring sites are empty. We estimate critical exponents theta, beta, nu||, and nu perpendicular, characterizing, respectively, the density of active particles in time, the order parameter, the temporal and spatial correlation lengths near the critical point, and the results are confirmed by off-critical scaling and finite size scaling. The order parameter exponent beta on a checkerboard fractal appears to lie between the one-dimensional (1D) value and two-dimensional (2D) value of the CLG model, while that on a Sierpinski gasket lies between the 1D and 2D values of the conserved threshold transfer process. Such a difference is manifested based on the intrinsic properties of the underlying fractal lattices.

Identification of differentially expressed genes in distinct skeletal muscles in cattle using cDNA microarray.

The 788-gene microarray was manufactured using selected elements from three different cDNA libraries in order to identify molecular processes that determine phenotypic characteristics between loin (M. longissimus thoracis) and round (M. semimembranosus) muscles. Microarray analyses identified 24 differentially expressed genes between the two muscles investigated. Five of the genes were verified by quantitative RT-PCR and three of them were mapped on bovine chromosomes using 5,000 rad bovine radiation hybrid (RH) panel. The map locations indicated that they were mapped in the same chromosomal regions where IMF and growth QTLs were located, suggesting that they are most possible positional candidate genes for the traits.

Unilateral Spinous Process Noncovering Hook Type Patient-specific Drill Template for Thoracic Pedicle Screw Fixation: A Pilot Clinical Trial and Template Classification.

STUDY DESIGN: Prospective clinical pilot study. OBJECTIVE: To confirm the accuracy of thoracic pedicle screw placement by using the unilateral spinous noncovering hook type patient-specific drill template (PSDT) made through rapid prototyping (RP) and to analyze previously reported PSDT designs and their characteristics. SUMMARY OF BACKGROUND DATA: Pedicle screw fixation is the most common form of the posterior instrumentation of the thoracic and lumbar spine. Various techniques have been introduced to improve pedicle screw placement. Among them PSDT with a preplanned trajectory has been considered a promising solution; however, we don't have consensus on proper character of the template. METHODS: Preoperative spiral three-dimensional (3D) computed tomography (CT) was performed on the thoracic spine. The images were stored in DICOM format and transferred to a workstation running MIMICS 17.0 software to generate a 3D reconstruction template for the desired thoracic vertebra. The accurate trajectory and screw diameter and length were calculated with UG Imageware 12.1. The guide template was sterilized and used intraoperatively to assist with the placement of thoracic pedicle screws. After all pedicle trajectory screws had been inserted. We reviewed 12 previous reports and classified them according to the shape and system of PSDT that met the inclusion criteria of the review. RESULTS: Ten screws were placed by using the PSDT without violating the single laminar cortex. There was no violation of the spinal canal or the cortex of pedicle on postoperative CT scans. The results of 13 PSDT types included in the current study suggested that there is no significant difference in accuracy between the PSDTs. CONCLUSION: The unilateral spinous process noncovering hook type PSDT made through RP provided an accurate trajectory for the thoracic vertebra, and the classification of PSDT in this study could be helpful for further studies. LEVEL OF EVIDENCE: 5.

Visualization of venous systems by time-of-flight magnetic resonance angiography.

BACKGROUND AND PURPOSE: Time-of-flight (TOF) imaging technique depicts strong signal from fresh unsaturated magnetization that moves fast into the imaging region, and TOF magnetic resonance angiography (MRA) visualizes the arterial system using saturation pulse and band. However, TOF MRA can visualize the venous system when the flow direction is reversed. METHODS AND PATIENTS: We consecutively enrolled patients between June 2002 and February 2003 with an internal jugular vein (IJV) and sigmoid sinus (SS) visualized by TOF MRA. Carotid Duplex ultrasonography was performed on all patients to check IJV flow directions. Gadolinium (Gd)-enhanced MRA and conventional digital subtraction angiography were performed in selected patients. RESULTS: The IJVs and SSs of eight patients (left = 7, right = 1) were observed by TOF MRA. In these 8 patients, Duplex ultrasonography confirmed a reversed direction in IJVs. Four of the patients underwent Gd-enhanced MRA, which showed proximal innominate vein steno-occlusion. CONCLUSIONS: This is the first description of IJV and SS visualization by TOF MRA. Clinical implications are discussed.

Distinct regions of p53 have a differential role in transcriptional activation and repression functions.

The wild type p53 tumor suppressor protein transactivates genes carrying p53 responsive elements and represses several TATA containing promoters. We report in vivo gene regulation assays where deletion of the N-terminal 75 residues (delta N75) results in loss of transactivation of p53CON and repression of an HPV 6 reporter. In contrast, removal of the C-terminal 75 (delta C75) amino acids resulted in a truncated protein capable of trans-activating p53CON but not able to repress the HPV 6 reporter. In vitro protein association assays revealed that the delta N75 protein, but not the delta C75 truncated protein, could oligomerize with the wild type p53 protein. Co-transfection assays with wild type p53 showed that the delta N75 mutant protein has a dominant negative effect on trans-activation function. However, it does not affect the ability of wild type p53 to repress transcription from the HPV 6 receptor. The delta C75 protein had no effect on the ability of the wild type p53 to activate p53CON or repress the HPV 6 reporter. These results suggest that distinct regions of p53 have a differential role in transcriptional activation and repression functions.