Optimizing 4D abdominal MRI: image denoising using an iterative back-projection approach.

4D-MRI is a promising tool for organ exploration, target delineation and treatment planning. Intra-scan motion artifacts may be greatly reduced by increasing the imaging frame rate. However, poor signal-to-noise ratios (SNR) are observed when increasing spatial and/or frame number per physiological cycle, in particular in the abdomen. In the current work, the proposed 4D-MRI method favored spatial resolution, frame number, isotropic voxels and large field-of-view (FOV) during MR-acquisition. The consequential SNR penalty in the reconstructed data is addressed retrospectively using an iterative back-projection (IBP) algorithm. Practically, after computing individual spatial 3D deformations present in the images using a deformable image registration (DIR) algorithm, each 3D image is individually enhanced by fusing several successive frames in its local temporal neighborood, these latter being likely to cover common independent informations. A tuning parameter allows one to freely readjust the balance between temporal resolution and precision of the 4D-MRI. The benefit of the method was quantitatively evaluated on the thorax of 6 mice under free breathing using a clinically acceptable duration. Improved 4D cardiac imaging was also shown in the heart of 1 mice. Obtained results are compared to theoretical expectations and discussed. The proposed implementation is easily parallelizable and optimized 4D-MRI could thereby be obtained with a clinically acceptable duration.

Three-dimensional ultrashort echo time (3D UTE) MRI of Achilles tendon at 4.7T MRI with comparison to conventional sequences in an experimental murine model of spondyloarthropathy.

BACKGROUND: Due to the very short T2 of its components, the normal anatomy of Achilles enthesis is impossible to define with "conventional" long echo time (TE) T2 sequences. However, this is a common site affected by rheumatologic disease. Early abnormalities related to inflammatory processes are impossible to detect in this location. PURPOSE: To assess the feasibility of a 3D-UTE (ultrashort echo time) sequence to evaluate normal and pathological Achilles entheses, determining both anterior fibrocartilaginous and posterior collagenic portions at 4.7T, in a rat model of spondyloarthropathy (SpA) with histological correlation. To assess whether this sequence detects SpA enthesopathy prior to long TE T2 sequences, enabling disease monitoring. STUDY TYPE: Prospective case-control study. ANIMAL MODEL: Twelve immunocompetent Wistar male rats imaged before (controls); the model was induced in eight rats (16 tendons) imaged at day 6, day 13, and day 21 with regular sacrifice for ex vivo imaging and histological correlation. FIELD STRENGTH: 4.7T Bruker Biospec Systems. 3D balanced steady-state free precession (bSSFP) and 3D-UTE sequences, performed at baseline (day 0, n = 12 animals / 24 tendons), day 6 (n = 8/16), 13 (n = 4/8), and day 21 (n = 2/4). ASSESSMENT: Visual analysis and signal intensity measurements (signal to noise ratio, SNR) of both bSSFP and UTE images were performed by two independent musculoskeletal radiologists at different locations of the Achilles enthesis and preinsertional area. STATISTICAL TESTS: Normal and pathological rat values were compared by Wilcoxon signed-rank tests, as well as interobserver differences. MRI findings were compared against histological data. RESULTS: The 3D-UTE sequence identified the anterior fibrocartilage and posterior collagenic areas of Achilles entheses in all cases. Visual analysis and signal intensity measurements distinguished SpA-affected entheses from healthy ones at days 6 and 13 (P = 0.002 and P = 0.006, respectively). Neither the normal anatomy of the enthesis nor its pathological pattern could be identified on T2 bSSFP sequences. DATA CONCLUSION: Unlike bSSFP T2 sequences, 3D-UTE sequences enable visualization of normal enthesis anatomy and early detection of abnormalities in pathological conditions. LEVEL OF EVIDENCE: 2 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2019;50:127-135.

In vivo MRI discrimination between live and lysed iron-labelled cells using balanced steady state free precession.

OBJECTIVES: The goal of this study was to evaluate the ability of balanced steady state free precession (b-SSFP) magnetic resonance imaging sequence to distinguish between live and lysed iron-labelled cells. METHODS: Human breast cancer cells were labelled with iron oxide nanoparticles. Cells were lysed using sonication. Imaging was performed at 3 T. The timing parameters for b-SSFP and the number of iron-labelled cells in samples were varied to optimise the b-SSFP signal difference between live and lysed iron-labelled cell samples. For in vivo experiments, cells were mixed with Matrigel and implanted into nude mice. Three mice implanted with live labelled cancer cells were irradiated to validate this method. RESULTS: Lysed iron-labelled cells have a significantly higher signal compared with live, intact iron-labelled cells in bSSFP images. The contrast between live and dead cells can be maximised by careful optimisation of timing parameters. A change in the b-SSFP signal was measured 6 days after irradiation, reflecting cell death in vivo. Histology confirmed the presence of dead cells in the implant. CONCLUSIONS: Our results show that the b-SSFP sequence can be optimised to allow for the discrimination of live iron-labelled cells and lysed iron-labelled cells in vitro and in vivo.

PulseNet USA standardized pulsed-field gel electrophoresis protocol for subtyping of Vibrio parahaemolyticus.

PulseNet is a national molecular subtyping network for foodborne disease surveillance composed of public health and food regulatory agencies. Participants employ molecular subtyping of foodborne pathogens using a standardized method of pulsed-field gel electrophoresis (PFGE) for conducting laboratory-based surveillance of foodborne pathogens. The PulseNet standardized PFGE protocols are developed through a comprehensive testing process. The reproducibility of the protocol undergoes an internal evaluation at the Centers for Disease Control and Prevention and an external evaluation in multiple PulseNet laboratories. Here we describe the development and evaluation of a rapid PFGE protocol for subtyping Vibrio parahaemolyticus for use in PulseNet activities. The protocol was derived from the existing standardized PulseNet protocols for Escherichia coli O157:H7 and Vibrio cholerae. An external evaluation of this protocol was undertaken in collaboration with three PulseNet USA participating public health laboratories. Comparative analysis of the PFGE fingerprints generated by each of these laboratories demonstrated that the protocol is both reliable and reproducible in the hands of multiple users.

Microglia used as vehicles for both inducible thymidine kinase gene therapy and MRI contrast agents for glioma therapy.

Microglia are phagocytic cells that are chemoattracted by brain tumors and can represent up to 70% of the tumor cell population. To get insight into gene therapy against glioma, we decided to take advantage of those microglia properties and to use those cells as vehicles to transport simultaneously a suicide gene (under the control of a heat-sensitive promoter) and contrast agents to localize them by magnetic resonance imaging before applying any therapeutic treatment. Thymidine kinase (TK) expression and its functionality after gancyclovir administration were investigated. After the heat shock (44 degrees C and 20 min), TK was expressed in 50% of the cells. However, after gancyclovir treatment, 90% of the cells died by apoptosis, showing an important bystander effect. Then, the cells were incubated with new lanthanide contrast agents to check both their potential toxicity and their MR properties. Results indicate that the nanoparticles did not induce any cell toxicity and yield a hypersignal on MR images at 4.7 T. These in vitro experiments indicate that microglia are good candidates as vectors in gene therapy against brain tumors. Finally, microglia containing gadolinium-grafted nanoparticles were injected in the close vicinity of C6 tumor, in a mouse. The hyperintensive signal obtained on in vivo images as well as its retention time show the potential of the novel contrast agents for cellular imaging.

Increase of Fas-induced apoptosis by inhibition of extracellular phosphorylation of Fas receptor in Jurkat cell line.

Apoptosis signalling through the Fas pathway requires several steps of aggregation of the Fas receptor in the membrane, including aggregation that may occur in the absence of Fas ligand. Association of Fas domains is determinant to signal transmission following Fas ligand binding to a specific domain. The domains involved in Fas aggregation are located in its extracellular region and contain three potential protein kinase C-binding motifs. We therefore studied the possibility that phosphorylation of the extracellular region of Fas might be implicated in the regulation of Fas-mediated apoptosis. Inhibition experiments of extracellular phosphorylation were performed in human Jurkat T leukemia cells with K252b, an impermeant protein-kinase inhibitor. Extracellular phosphorylation of Fas receptor was related to ecto-kinase, as assessed by the [gamma-(32)P] ATP labelling of Fas-116 kDa aggregates, suppressed by K252b inhibitor which significantly increased the sensitivity to Fas-mediated apoptosis. Ecto-PKC involvement was demonstrated by bisindolylmaleimide VIII, a selective inhibitor of protein kinase C which significantly increased both Fas aggregation in the membrane and Fas-mediated apoptosis and by the addition of the PKC pseudo-substrate 19-36 which inhibited the phosphorylation of 116 kDa Fas aggregates. These data support a role for Fas phosphorylation in the decreased sensitivity to apoptosis in the Jurkat T leukemia cell line.

PulseNet USA: a five-year update.

PulseNet USA is the molecular surveillance network for foodborne infections in the United States. Since its inception in 1996, it has been instrumental in detection, investigation and control of numerous outbreaks caused by Shiga toxin-producing Escherichia coli O157:[H7] (STEC O157), Salmonella enterica, Listeria monocytogenes, Shigella spp., and Campylobacter. This paper describes the current status of the network, including the methodologies used and its future possibilities. The currently preferred subtyping method in the network is pulsed-field gel electrophoresis (PFGE), a proven highly discriminatory molecular subtyping method. New simpler sequencebased subtyping methods are under development and validation to complement and eventually replace PFGE. PulseNet is essentially a cluster detection network, but the data in the system will now also be used in attribution analyses of sporadic infections. The PulseNet platform will also be used as a primary tool in preparedness and response to acts of food bioterrorism.

Development and validation of a PulseNet standardized pulsed-field gel electrophoresis protocol for subtyping of Vibrio cholerae.

PulseNet is a network that utilizes standardized pulsed-field gel electrophoresis (PFGE) protocols with the purpose of conducting laboratory-based surveillance of foodborne pathogens. PulseNet standardized PFGE protocols are subject to rigorous testing during the developmental phase and careful evaluation during a validation process assessing its robustness and reproducibility in different laboratories. Here we describe the development and validation of a rapid PFGE protocol for subtyping Vibrio cholerae for use in PulseNet International activities. While the protocol was derived from the existing PulseNet protocol for Escherichia coli O157, various aspects of this protocol were optimized for use with V. cholerae, most notably a change of the primary and secondary restriction enzyme to SfiI and NotI, respectively, and the use of a two-block electrophoresis program. External validation of this protocol was undertaken through a collaboration between three PulseNet Asia Pacific laboratories (Public Health Laboratory Centre, Hong Kong, National Institute of Infectious Diseases, Japan, and International Center for Diarrhoeal Diseases Research-Bangladesh) and PulseNet USA. Comparison of PFGE patterns generated by each of the participating laboratories demonstrated that the protocol is robust and reproducible.

[Small cell lung cancer and cancer-associated retinopathy]. / Carcinoma broncogénico microcítico y retinopatía asociada a cáncer.

We report the case of a patient who presented with cancer-associated retinopathy and small cell carcinoma of the lung, which was treated surgically because the initial diagnostic biopsy finding was squamous cell carcinoma. The patient then underwent chemotherapy and radiation therapy. We discuss the characteristics and pathogenesis of this paraneoplastic syndrome as well as its association with the lung tumor's aberrant production of a protein that competes with retinal recoverin at the photoreceptors of the retinal cone.

Prostaglandins mediate lipopolysaccharide effects upon cholecystokinin and neurotensin phenotypes in neuroendocrine corticotropin-releasing hormone neurons: in situ hybridization evidence in the rat.

Intraperitoneal injection of the endotoxin lipopolysaccharide produces an inflammation accompanied by immune system activation and secretion of cytokines that stimulate the hypothalamo-pituitary-adrenal (HPA) axis to release the anti-inflammatory corticosterone. Upstream in HPA axis are neuroendocrine corticotropin-releasing hormone neurons in the paraventricular nucleus whose multipeptidergic phenotype changes during inflammation: coexisting corticotropin-releasing hormone and cholecystokinin mRNAs are up-regulated whereas neurotensin mRNA expression is induced de novo. These changes may be mediated by prostaglandins released from perivascular and microglial cells in response to circulating cytokines. We examined by quantitative in situ hybridization histochemistry whether blockade of prostaglandin synthesis by indomethacin alters phenotypic expression in paraventricular nucleus neurons after lipopolysaccharide. Because indomethacin also elevated circulating corticosterone, animals were adrenalectomized and corticosterone replaced. Results showed that i.p. indomethacin administration suppressed lipopolysaccharide effects in a phenotype non-specific manner: one injection was sufficient to prevent both the increase in corticotropin-releasing hormone and cholecystokinin mRNAs expression and the induction of neurotensin mRNA expression. Therefore, neuroendocrine corticotropin-releasing hormone neurons with different peptidergic phenotypes appear to respond as a whole in the acute phase response to systemic infection.

Rapid pulsed-field gel electrophoresis protocol for subtyping of Campylobacter jejuni.

We developed a rapid pulsed-field gel electrophoresis (PFGE) protocol for subtyping Campylobacter isolates based on the standardized protocols used by PulseNet laboratories for the subtyping of other food-borne bacterial pathogens. Various combinations of buffers, reagents, reaction conditions (e.g., cell suspension concentration, lysis time, lysis temperature, and restriction enzyme concentration), and electrophoretic parameters were evaluated in an effort to devise a protocol that is simple, rapid, and robust. PFGE analysis of Campylobacter isolates can be completed in 24 to 30 h using this protocol, whereas the most widely used current protocols require 3 to 4 days to complete. Comparison of PFGE patterns obtained in six laboratories showed that subtyping results obtained using this protocol are highly reproducible.

Evaluation of pulsed-field gel electrophoresis in epidemiological investigations of meningococcal disease outbreaks caused by Neisseria meningitidis serogroup C.

Since 1990, the frequency of Neisseria meningitidis serogroup C (NMSC) outbreaks in the United States has increased. Based on multilocus enzyme electrophoresis (MEE), the current molecular subtyping standard, most of the NMSC outbreaks have been caused by isolates of several closely related electrophoretic types (ETs) within the ET-37 complex. We chose 66 isolates from four well-described NMSC outbreaks that occurred in the United States from 1993 to 1995 to evaluate the potential of pulsed-field gel electrophoresis (PFGE) to identify outbreak-related isolates specific for each of the four outbreaks and to differentiate between them and 50 sporadic isolates collected during the outbreak investigations or through active laboratory-based surveillance from 1989 to 1996. We tested all isolates collected during the outbreak investigations by four other molecular subtyping methods: MEE, ribotyping (ClaI), random amplified polymorphic DNA assay (two primers), and serotyping and serosubtyping. Among the 116 isolates, we observed 11 clusters of 39 NheI PFGE patterns. Excellent correlation between the PFGE and the epidemiological data was observed, with an overall sensitivity of 85% and specificity of 71% at the 95% pattern relatedness breakpoint using either 1.5 or 1.0% tolerance. For all four analyzed outbreaks, PFGE would have given public health officials additional support in declaring an outbreak and making appropriate public health decisions.

Is the mineralocorticoid receptor in Brown Norway rats constitutively active?

In a previous study using corticosterone treatment of adrenalectomized rats, we hypothesized that mineralocorticoid receptor (MR)-related mechanisms are constitutively active and that glucocorticoid receptor (GR)-mediated mechanisms are more efficient in Brown Norway rats compared to Fischer 344 (F344) rats. In order to discriminate the mineralocorticoid from the glucocorticoid actions exerted by corticosterone, F344 and Brown Norway adrenalectomized rats were treated with increasing doses (1, 5 and 25 microg/ml of drinking water) of deoxycorticosterone (DOC, MR-specific ligand) or RU 28362 (GR-specific ligand). These rats were compared with long-term adrenalectomized (ADX) untreated rats and sham-ADX rats. This study confirms our previous results, notably the lack of effect of ADX on body weight and fluid intake in Brown Norway rats. Moreover, DOC treatment had no effect in Brown Norway rats whereas the higher dose restored fluid intake of the F344 ADX group to sham values. These results support the hypothesis of a constitutive activation of the MR and therefore the insensitivity of this receptor to its ligand in Brown Norway rats. Alternatively, RU 28362 treatment induced greater weight loss, decrease in food intake, anxiolysis, thymus involution, and decrease in plasma transcortin concentration and pituitary corticosteroid receptor densities in Brown Norway rats than in F344 rats, which is consistent with greater efficiency of GR mechanisms in Brown Norway rats than in F344 rats. Therefore, these strains are of great utility to disentangle MR and GR effects on complex phenotypes.

Ceftriaxone-resistant salmonella infection acquired by a child from cattle.

BACKGROUND: The emergence of resistance to antimicrobial agents within the salmonellae is a worldwide problem that has been associated with the use of antibiotics in livestock. Resistance to ceftriaxone and the fluoroquinolones, which are used to treat invasive salmonella infections, is rare in the United States. We analyzed the molecular characteristics of a ceftriaxone-resistant strain of Salmonella enterica serotype typhimurium isolated from a 12-year-old boy with fever, abdominal pain, and diarrhea. METHODS: We used pulsed-field gel electrophoresis and analysis of plasmids and beta-lactamases to compare the ceftriaxone-resistant S. enterica serotype typhimurium from the child with four isolates of this strain obtained from cattle during a local outbreak of salmonellosis. RESULTS: The ceftriaxone-resistant isolate from the child was indistinguishable from one of the isolates from cattle, which was also resistant to ceftriaxone. Both ceftriaxone-resistant isolates were resistant to 13 antimicrobial agents; all but one of the resistance determinants were on a conjugative plasmid of 160 kb that encoded the functional group 1 beta-lactamase CMY-2. Both ceftriaxone-resistant isolates were closely related to the three other salmonella isolates obtained from cattle, all of which were susceptible to ceftriaxone. CONCLUSIONS: This study provides additional evidence that antibiotic-resistant strains of salmonella in the United States evolve primarily in livestock. Resistance to ceftriaxone, the drug of choice for invasive salmonella disease, is a public health concern, especially with respect to children, since fluoroquinolones, which can also be used to treat this disease, are not approved for use in children.

Emergence of domestically acquired ceftriaxone-resistant Salmonella infections associated with AmpC beta-lactamase.

CONTEXT: Ceftriaxone, an expanded-spectrum cephalosporin, is an antimicrobial agent commonly used to treat severe Salmonella infections, especially in children. Ceftriaxone-resistant Salmonella infections have recently been reported in the United States, but the extent of the problem is unknown. OBJECTIVES: To summarize national surveillance data for ceftriaxone-resistant Salmonella infections in the United States and to describe mechanisms of resistance. DESIGN AND SETTING: Case series and laboratory evaluation of human isolates submitted to the Centers for Disease Control and Prevention from 17 state and community health departments participating in the National Antimicrobial Resistance Monitoring System (NARMS) for enteric bacteria between 1996 and 1998. PATIENTS: Patients with ceftriaxone-resistant Salmonella infections between 1996 and 1998 were interviewed and isolates with decreased ceftriaxone susceptibility were further characterized. MAIN OUTCOME MEASURES: Exposures and illness outcomes, mechanisms of resistance. RESULTS: The prevalence of ceftriaxone-resistant Salmonella was 0.1% (1 of 1326) in 1996, 0.4% (5 of 1301) in 1997, and 0.5% (7 of 1466) in 1998. Ten (77%) of the 13 patients with ceftriaxone-resistant infections were aged 18 years or younger. The patients lived in 8 states (California, Colorado, Kansas, Massachusetts, Maryland, Minnesota, New York, and Oregon). Nine (82%) of 11 patients interviewed did not take antimicrobial agents and 10 (91%) did not travel outside the United States before illness onset. Twelve of the 15 Salmonella isolates with ceftriaxone minimum inhibitory concentrations of 16 microg/mL or higher were serotype Typhimurium but these isolates had different pulsed-field gel electrophoresis patterns. Thirteen of these 15 isolates collected between 1996 and 1998 were positive for a 631-base pair polymerase chain reaction product obtained by using primers specific for the ampC gene of Citrobacter freundii. CONCLUSIONS: Domestically acquired ceftriaxone-resistant Salmonella has emerged in the United States. Most ceftriaxone-resistant Salmonella isolates had similar AmpC plasmid-mediated resistance.

Evaluation of the chick embryo for the determination of relative virulence of Neisseria meningitidis.

The chick embryo model was evaluated as a method to compare virulence between selected strains of Neisseria meningitidis. Inoculation of 13-day-chick embryos via the egg yolk distinguished strains having an LD50 of 10(3) colony forming units (CFU) or greater (low virulence) from those having an LD50 of approximately 10(1) or less (high virulence). A strain of serogroup B and a spontaneous nonpiliated strain of group C were found to be of relatively high virulence while a strain of N. lactamica, a serogroup A carrier strain, and certain nongroupable strains were found to be of low virulence. Strains having an LD50 of 10(2) were not differentiated from either of these. Alternatively, inoculation of the chorioallantoic membrane (CAM) of 9-day-old chick embryos statistically differentiated most strains of N. meningitidis although inoculation via this route was less sensitive.