Comparison of contrast-enhanced ultrasonography and MRI results obtained by expert and novice radiologists indicating short-term response after transarterial chemoembolisation for hepatocellular carcinoma.

AIM: To evaluate inter-reader agreement between novice and expert radiologists in assessing contrast-enhanced ultrasonography (CEUS) and magnetic resonance imaging (MRI) images for detecting viable tumours with different sizes after conventional transarterial chemoembolisation (cTACE). MATERIALS AND METHODS: This prospective study included patients who had less than five hepatomas and who underwent cTACE. Hepatomas with one or two feeding arteries were selected as target lesions. CEUS and MRI were performed within 1 week after cTACE to evaluate viable tumours. RESULTS: The expert group had higher kappa values in evaluating all tumour sizes via CEUS compared with MRI. The novice group had similar kappa values. In patients with tumours measuring ≤3 cm, the expert group had higher kappa values in reading CEUS compared with MRI images; however, in the novice group, the kappa value was lower in evaluating CEUS compared with MRI images. In patients with tumours measuring >3 cm, the expert and novice groups had good to excellent kappa values. The confidence level of the two groups in reading MRI images was high; however, the novice group had a lower confidence level. CONCLUSION: CEUS is a convenient, cost-effective, and easy to apply imaging tool that can help interventionists perform early detection of viable hepatocellular carcinoma post-TACE. It has a higher inter-rater agreement in interpreting CEUS images compared with MRI images among expert radiologists even when they are extremely familiar with post-cTACE MRI images. In novice radiologists, there may be a learning curve to achieve good consistency in CEUS interpretation.

A case of GNE myopathy mimicking hereditary motor neuropathy.

A 36-year-old woman who presented with upper limb distal weakness since the age of 15 years, with gradual progression to the lower limbs, is reported. Hereditary motor neuropathy was initially suspected based on distal weakness and hyporeflexia; however, whole exome sequencing accidentally revealed a compound heterozygous variant in the GNE gene, and ultrasound revealed increased homogeneous echogenicity in the involved muscles, which is characteristic of myopathic changes. Muscle magnetic resonance imaging revealed fatty infiltration in all limb muscles, sparing the triceps brachii, vastus lateralis and vastus medialis. Muscle biopsy revealed intracytoplasmic rimmed vacuole, supporting the diagnosis of GNE myopathy.

A critical role for lymphotoxin-beta receptor in the development of diabetes in nonobese diabetic mice.

To assess the role of lymphotoxin-beta receptor (LTbetaR) in diabetes pathogenesis, we expressed an LTbetaR-Fc fusion protein in nonobese diabetic (NOD) mice. The fusion protein was expressed in the embryo, reached high levels for the first 2 wk after birth, and then declined progressively with age. High expression of LTbetaR-Fc blocked diabetes development but not insulitis. After the decline in chimeric protein concentration, mice became diabetic with kinetics similar to the controls. Early expression of fusion protein resulted in disrupted splenic architecture. However, primary follicles and follicular dendritic cells, but not marginal zones, developed in aged mice. Hence, LTbetaR signaling is required for diabetes development and regulates follicular and marginal zone structures via qualitatively or quantitatively distinct mechanisms.

Occurrence of haloacetic acids (HAAs) and trihalomethanes (THMs) in drinking water of Taiwan.

In this study, water samples were collected from 86 water treatment plants for analysis of haloacetic acids (HAAs) and trihalomethanes (THMs) from February to March, 2007 and from July to August, 2007. Both seasonal and geographical variations of disinfection by-products (DBPs) in drinking water of Taiwan were presented. The results showed that the five HAA concentrations (HAA5) were 1.0-38.9 microg/L in the winter and 0.2-46.7 microg/L in the summer; and the total THMs were ND-99.4 microg/L in the winter and ND-133.2 microg/L in the summer. For samples taken from the main Taiwan island, dichloroacetic acid (29.4-31.7%) and trichloroacetic acid (25.3-27.6%) were the two major HAA species, and trichloromethane was the major THM species (49.9-62.2%) in finished water. For water treatment plants located on the offshore islands outside of Taiwan, high bromide concentration was found in raw water, and higher percentage of brominated THMs and HAAs were formed in the overall formation. A statistically significant (P < 0.005) logarithmic linear regression model was found to be useful to describe the correlations between TTHM and HAA5 or nine HAAs (HAA5 = 1.219 x TTHM (0.754), R(2) = 0.658; HAA9 = 1.824 x TTHM (0.735), R(2) = 0.678). No apparent difference was observed for DBPs concentrations between finished water and distribution samples in this study.

Involvement of Gas7 along the ERK1/2 MAP kinase and SOX9 pathway in chondrogenesis of human marrow-derived mesenchymal stem cells.

OBJECTIVE: The growth-arrest-specific protein, Gas7, has been shown to be involved in reorganization of the cytoskeleton and for inducing changes in cell shape during cell differentiation. The goals of this study were to investigate the novel role of human Gas7 (hGas7) in chondrogenic differentiation of human mesenchymal stem cells (hMSCs) and to identify the relationship between hGas7, extracellular signal-regulated kinase (ERK1/2) and SOX9 in the chondrogenic pathway. METHODS: Bone marrow-derived hMSCs were induced to undergo chondrogenic differentiation with transforming growth factor-beta1 (TGF-beta1) in an aggregate culture system. The expression of hGas7 and SOX9 and phosphorylation of ERK1/2 at multiple time points were investigated. Chondrogenic capacity was evaluated by the size of aggregates, by glycosaminoglycan content, and by type II collagen and proteoglycan deposition after interfering with expression of hGas7, ERK1/2 or SOX9. To delineate the functional role of these genes in chondrogenesis, inhibition of individual gene's expression in hMSCs, by antisense oligonucleotides or interference RNA (siRNA), and the effect on chondrogenic differentiation were also investigated. RESULTS: Treatment of hMSCs with TGF-beta1 resulted in a transient up-regulation of hGas7b, one of the hGas7 isoforms (day 3-day 5), a transient phosphorylation of ERK1/2 (0.5-4 h) and an up-regulation of SOX9 (2 h to day 14). Transient expression of hGas7b was also detected in hMSCs by reverse transcription-polymerase chain reaction at day 2 and day 3 following TGF-beta1 treatment. Interference with hGas7b production by hGas7b-specific antisense oligonucleotide or inhibition of p-ERK with PD98059, a specific inhibitor of ERK signaling pathway, or interference with SOX9 production by SOX9 siRNA all caused adverse effects of chondrogenic differentiation of hMSCs. Meanwhile, inhibition of p-ERK or SOX9 both blocked the expression of hGas7b. However, the p-ERK and SOX9 pathway was not affected by inhibition of hGas7b. CONCLUSION: These results provide evidence that the transient expression of hGas7b, regulated by activation of ERK1/2 and SOX9 pathway, is essential for chondrogenic differentiation of hMSCs.

Painful neuropathy with skin denervation after prolonged use of linezolid.

The prolonged use of linezolid, a new antibiotic against drug-resistant gram-positive pathogens, might cause painful neuropathy. This finding raises the possibility that small-diameter sensory nerves in the skin, which are responsible for transmitting nociceptive information, might be affected. We report a 53-year-old female who developed pure small-fibre painful neuropathy (visual analogue scale, VAS = 82 on 0-100 scale) with marked skin denervation in the leg (epidermal nerve density, END = 2.32 fibres/mm, norm <5.88 fibres/mm) and significant elevation of the warm threshold in the foot (40.0 degrees C, norm <39.4 degrees C) after the use of linezolid for 6 months. Eight months after the discontinuation of linezolid, the skin became fully reinnervated (END = 9.04 fibres/mm), with disappearance of neuropathic pain (VAS = 0) and normalisation of the warm threshold (36.3 degrees C). Nerve conduction studies for large-diameter motor and sensory nerves were normal. This report documents a pure small-fibre sensory neuropathy after prolonged use of linezolid, and the relationship between skin innervation and corresponding neuropathic pain.

Metabolic characterization of serum from mice challenged with Orientia tsutsugamushi-infected mites.

Scrub typhus is an acute zoonosis caused by the obligate intracellular Gram-negative bacterium Orientia tsutsugamushi. To better understand the host response elicited by natural infection by chigger feeding, ICR mice were infected by Leptotrombidium chiangraiensis (Lc1) chiggers, and the metabolic profiles of their serum were examined over several time points after initiation of feeding. ICR mice were infected by either naive Lc1 chiggers (i.e. not infected by O. tsutsugamushi, NLc1) or O. tsutsugamushi-infected Lc1 chiggers (OLc1). Serum was collected from both groups of mice at 6 hours and 10 days after initiation of feeding. Metabolites were extracted from the serum and analysed by ultra performance liquid chromatography-tandem mass spectrometry. The resulting ion/chromatographic features were matched to a library of chemical standards for identification and quantification. Biochemicals that differed significantly between the experimental groups were identified using Welch's two-sample t tests; p ≤ 0.05 was considered statistically significant. A number of biochemicals linked to immune function were found to be significantly altered between mice infected by the NLc1 and OLc1 chiggers, including itaconate, kynurenine and histamine. Several metabolites linked to energy production were also found to be altered in the animals. In addition lipid and carbohydrate metabolism, bile acid and phospholipid homeostasis, and nucleotide metabolism were also found to be different in these two groups of mice. Markers of stress and food intake were also significantly altered. Global untargeted metabolomic characterization revealed significant differences in the biochemical profiles of mice infected by the NLc1 versus OLc1 chiggers. These findings provide an important platform for further investigation of the host responses associated with chigger-borne O. tsutsugamushi infections.

Serological evidence of high Leptospira exposure among indigenous people (Orang Asli) in Peninsular Malaysia using a recombinant antigen-based ELISA.

The lifestyles of the indigenous people (Orang Asli) of Peninsular Malaysia who traditionally live close to the forest, put them at higher risk of exposure to zoonotic diseases. Leptospirosis has recently emerged as one of the most important diseases of public health concern. Here, we aimed to obtain a baseline data on the level of Leptospira exposure among the 107 Orang Asli volunteers using a recombinant antigen-based ELISA, previously shown to have sensitivity of ~90.0% in comparison to the microscopic agglutination test (MAT). Among the Orang Asli volunteers in this study, 60.7% had IgM against Leptospira and 57.9% were antiLeptospira IgG positive. Of these seropositive individuals, 29.9% had both anti-Leptospira IgM and IgG antibodies. Age was found to be a significant predictor for exposure to Leptospira (P < 0.05) with the younger Orang Asli population more likely to be tested positive for antiLeptospira IgM. The finding of high Leptospira exposure among the Orang Asli volunteers could be due to their socio-economic practices and dependency on the forest for their livelihood. The rapid and sensitive recombinant antigen-based ELISA used in the study, could possibly complement MAT for the epidemiological surveillance of leptospirosis, especially among the underserved populations.

Physiological and pathological characterization of capsaicin-induced reversible nerve degeneration and hyperalgesia.

BACKGROUND: The study aimed to investigate the physiology, psychophysics, pathology and their relationship in reversible nociceptive nerve degeneration, and the physiology of acute hyperalgesia. METHODS: We enrolled 15 normal subjects to investigate intraepidermal nerve fibre (IENF) density, contact heat-evoked potential (CHEP) and thermal thresholds during the capsaicin-induced skin nerve degeneration-regeneration; and CHEP and thermal thresholds at capsaicin-induced acute hyperalgesia. RESULTS: After 2-week capsaicin treatment, IENF density of skin was markedly reduced with reduced amplitude and prolonged latency of CHEP, and increased warm and heat pain thresholds. The time courses of skin nerve regeneration and reversal of physiology and psychophysics were different: IENF density was still lower at 10 weeks after capsaicin treatment than that at baseline, whereas CHEP amplitude and warm threshold became normalized within 3 weeks after capsaicin treatment. Although CHEP amplitude and IENF density were best correlated in a multiple linear regression model, a one-phase exponential association model showed better fit than a simple linear one, that is in the regeneration phase, the slope of the regression line between CHEP amplitude and IENF density was steeper in the subgroup with lower IENF densities than in the one with higher IENF densities. During capsaicin-induced hyperalgesia, recordable rate of CHEP to 43 °C heat stimulation was higher with enhanced CHEP amplitude and pain perception compared to baseline. CONCLUSIONS: There were differential restoration of IENF density, CHEP and thermal thresholds, and changed CHEP-IENF relationships during skin reinnervation. CHEP can be a physiological signature of acute hyperalgesia. SIGNIFICANCE: These observations suggested the relationship between nociceptive nerve terminals and brain responses to thermal stimuli changed during different degree of skin denervation, and CHEP to low-intensity heat stimulus can reflect the physiology of hyperalgesia.

Human cytomegalovirus-stimulated peripheral blood mononuclear cells induce HIV-1 replication via a tumor necrosis factor-alpha-mediated mechanism.

Human cytomegalovirus (HCMV) is a potential cofactor in HIV-1 infection. To investigate the mechanism whereby HCMV promotes HIV-1 replication, a PBMC coculture assay which measures HIV-1 p24 antigen release was used as an index of viral replication. HCMV-stimulated PBMC were capable of inducing HIV-1 replication in cocultures with acutely infected PBMC; however, this occurred only when the PBMC were from HCMV-seropositive donors (598 +/- 207 versus 27 +/- 10 pg/ml p24 antigen with PBMC from HCMV-seronegative donors on day 6 of coculture). Upon stimulation with HCMV, PBMC obtained exclusively from HCMV-seropositive donors released tumor necrosis factor (TNF)-alpha (270 +/- 79 pg/ml at 18 h of culture). Monoclonal antibodies to TNF-alpha blocked the activity of HCMV-stimulated PBMC in cocultures both with acutely HIV-1-infected PBMC and with the chronically infected promonocytic line U1. Also, treatment of HCMV-stimulated PBMC with pentoxifylline, an inhibitor of TNF-alpha mRNA, markedly reduced HIV-1 replication in cocultures both with acutely and chronically infected cells. These results indicate that TNF-alpha is a key mediator of HIV-1 replication induced by HCMV-stimulated PBMC and support the concept that this cytokine plays an important role in the pathogenesis of HIV-1 infection.

Effects of transforming growth factor-beta on murine astrocyte glutamine synthetase activity. Implications in neuronal injury.

Cytokines have been implicated in the pathogenesis of a number of brain diseases in which neurological dysfunction has been attributed to a change in amino acid neurotransmitter metabolism. In the present in vitro study, we investigated the effects of cytokines on astrocyte glutamine synthetase (GS) activity and subsequently on N-methyl-D-aspartate (NMDA) receptor-mediated neurotoxicity. Proinflammatory cytokines IL-1 alpha, IL-1 beta, and IL-6 at a concentration of 20 ng/ml did not affect GS activity; however, tumor necrosis factor-alpha inhibited this activity by 20% in mixed neuronal/astrocyte cultures. Treatment for 24 h with transforming growth factor (TGF)-beta 1 or -beta 2 inhibited up to 60% GS activity. TGF-beta 2 also inhibited GS in enriched astrocyte cultures with an ED50 of 10 pg/ml. Antibodies specific to TGF-beta 2 blocked this effect. Treatment of astrocytes with TGF-beta 2 (250 pg/ml) resulted in markedly dilated rough endoplasmic reticulum. Since astrocyte GS may play a protective role in NMDA receptor-mediated neurotoxicity, we treated mixed neuronal/astrocyte cultures with TGF-beta 2 (250 pg/ml) and found a threefold potentiation of NMDA receptor-mediated neurotoxicity. These data suggest that TGF-beta impairs astrocyte GS function and enhances neurotoxicity, thus providing insight into understanding one mechanism of cytokine-mediated central nervous system disease.

Transmission enhancement through a trench-surrounded nano metallic slit by bump reflectors.

We propose using a pair of bumps bordering the conventional trench-surrounded metal nano slit in order to confine the surface waves and further enhance the slit transmission. The bump height of 1.mum is larger than the depth of penetration on air side of the surface waves. The reflectivity of such bumps is larger than 95%. A very large slit transmission, which is 50% of the energy of the incident beam impinging on the entire size 13.mum of the trench-surrounded slit structure, is obtained through the metallic slit of 50nm width and 400nm depth. The bumps enhance the transmission by 1.75 fold.

Posttranslational translocation of influenza virus hemagglutinin across microsomal membranes.

The biosynthesis of influenza virus hemagglutinin (HA) and its translocation across microsomal membranes were studied in a mammalian cell-free system. All forms of HA could be cotranslationally translocated with high efficiency. However, only truncated forms of HA were translocated after protein synthesis has been terminated. The efficiency of this posttranslational translocation was dependent on the extent of the truncation. Posttranslational translocation was ribosome dependent and occurred only in the presence of a functional N-terminal signal sequence. The molecular mechanism of protein targeting and translocation across the membrane of the endoplasmic reticulum is discussed.

Cross-resistance to UV radiation of a cisplatin-resistant human cell line: overexpression of cellular factors that recognize UV-modified DNA.

A human cell line selected for cisplatin resistance (CPR) was irradiated with UV light and showed cross-resistance to UV light. Applying a modified chloramphenicol acetyltransferase assay, we observed that CPR cells acquired enhanced host cell reactivation of a transfected plasmid carrying UV damage. Gel mobility shift analysis indicated that two nuclear factors that recognize UV-modified DNA were overexpressed in CPR cells. In addition, factors that bind UV-modified DNA were independent from the factors that bind cisplatin-modified DNA. The significance of the identified binding factors, possibly DNA repair enzymes, is discussed.

Enhanced host cell reactivation of damaged plasmid DNA in HeLa cells resistant to cis-diamminedichloroplatinum(II).

Human HeLa cells resistant to cisplatin were established by stepwise selection. The selected cells showed a 15- to 20-fold cisplatin resistance (CPR) at the dose level resulting in 50% inhibition. These cells were cross-resistant to mitomycin C, melphalan, and ethyl methanesulfonate but not to Adriamycin, colchicine, or vinblastine. The expression of cisplatin-damaged plasmid DNA carrying the bacterial chloramphenicol acetyltransferase (CAT) gene after its transfection into CPR cells was enhanced by approximately 3-fold. This did not correlate with the degree of CPR. However, the development of the CPR phenotype paralleled the enhanced CAT activity. The addition of aphidicolin (an inhibitor of DNA alpha-polymerase) to CPR cells effectively diminished the enhanced CAT activity and CPR. These studies have identified an enhanced host cell reactivation of the damaged plasmid in the acquisition of CPR, suggesting that DNA repair is a potential mechanism for the development of CPR phenotype in human cells.

Ionic and ligand-specific effects on the DNA binding of progesterone receptor bound to the synthetic progestin R5020 and the antiprogestin RU486.

Antiprogestin and other antihormones are valuable therapeutic agents in hormone-responsive cancers. A fundamental mechanism in the action of antiprogestin is its binding to PR,3 an intracellular protein that mediates the action of progesterone by direct interaction at the regulatory sites of responsive genes. To elucidate the mechanism of action of PR bound to agonistic and antagonistic ligands, we determined the binding affinity of rabbit uterine PR bound to R5020 and RU486, respectively, with different DNA sequences. We used 2 recombinant plasmid DNAs, pDHf14 and pDHf2, with 60- and 23-base pair inserts of potential Z-DNA-forming (dA-dC)n.(dG-dT)n sequences, respectively, parental plasmid pDPL6 with no insert, calf thymus DNA, and several synthetic polynucleotides in this study. The concentration of DNA required to elute 50% of the receptor bound to DNA-cellulose (EC50) was used as a measure of the relative binding affinity of the receptor for DNA. EC50 values of plasmids pDHf14 and pDHf2 were 1.2 +/- 0.5 (SD) and 2.5 +/- 0.6 micrograms/ml, respectively, for PR bound to R5020. In contrast, EC50 for control plasmid was 350 micrograms/ml. PR.R5020 had lower affinity for calf thymus DNA and polynucleotides compared with pDHf2 and pDHf14. Receptors complexed with the antiprogestin RU486 had lower affinity for the plasmids; EC50 values were 2.4 +/- 0.4 and 10 +/- 1 microgram/ml, respectively, for pDHf14 and pDHf2. This ligand-specific difference in DNA binding was amplified by the presence of 5 mM Mg2+ or Ca2+. The relative binding affinity of PR.R5020 to pDHf14 was 6- and 7-fold higher than that of PR.RU486, in the presence of 5 mM Mg2+ and Ca2+, respectively. These results show that PR.RU486 has lower binding affinity for specific DNA sequences than PR.R5020, but the binding affinity of both receptors is in a range that cannot preclude competitive interactions at the DNA recognition site. The effects of Mg2+ and Ca2+ on PR binding to DNA further suggest that these cations could affect PR recognition of DNA in a ligand-specific manner.

Identification, cloning, and expression of potential diagnostic markers for Q fever.

The clinical diagnosis of Q fever is difficult. Whole cell antigens are currently used in several serological methods, but antigens are limited due to the hazardous nature of Coxiella burnetii cultivation. In this report, we described the method of detecting immunodominant antigens of C. burnetii by using proteomic techniques with patient sera, and cloning and expressing the selected antigens using a novel vector known for its ease of expression, purification, and downstream application.

Proteomic analysis of Rickettsia prowazekii.

Rickettsia prowazekii is an obligate intracellular gram-negative bacterium. Comparative proteomics study of a virulent strain (Breinl) versus an avirulent strain (Madrid E) was performed using an integrated liquid chromatography and mass spectrometer. About 30% of predicted proteins were detected and identified. Among the detected proteins, more than 30 proteins were of unknown function in both strains. Although several proteins were detected in only one strain, the overall distribution of detected proteins in different COGs (clusters of orthologs groups) was very similar between the two strains. Functional analysis of differentially expressed proteins, either qualitatively or quantitatively, may lead to the discovery of pathogenesis-related factors.

Inhibition of nitric oxide can ameliorate apoptosis and modulate matrix protein gene expression in bacteria infected chondrocytes in vitro.

Bacterial infection stimulates nitric oxide (NO) production in chondrocytes. However, the role of NO in chondrocyte apoptosis after infection remains unclear. The purpose of the study was to test if inhibition of NO could ameliorate apoptosis and modulate matrix protein gene expression in bacteria-infected chondrocytes. It was shown that pre-treating chondrocytes with L-NAME (1 mM) significantly decreased the release of NO (from 72 to 14 microM) and the extent of apoptosis (from 52.9% to 18.9%). Pre-treatment with L-NAME also counteracted the bacteria-induced downregulation of Type II collagen (from 26% to 79%) and aggrecan (from 63% to 105%) mRNA levels. Inhibition of NO after the induction of infection could not decrease the extent of apoptosis and modulate matrix protein gene expression. The results of this study support the hypothesis that NO has an important role in bacteria-induced chondrocyte apoptosis. Pre-treatment but not post-treatment could ameliorate the extent of apoptosis and reestablish the cartilage matrix protein gene expression. This study suggests that in addition to NO, other mechanisms may be responsible for the sustained destruction of articular cartilage in the post-infectious arthropathy.

Modulation of human microglial cell superoxide production by cytokines.

Reactive oxygen intermediates (e.g., superoxide [O2-]) generated by microglia may play a role in host defense and injury within the central nervous system. We investigated the effect of cytokines on human microglial cell O2- production on stimulation with phorbol myristate acetate. Priming of microglial cell cultures with interferon-gamma or tumor necrosis factor-alpha resulted in a dose- and time-dependent enhancement of O2- production. The priming effects of these cytokines were mediated through a protein kinase C signal transduction pathway. In contrast, astrocytes did not generate detectable O2- on phorbol myristate acetate stimulation. Treatment of microglia with transforming growth factor-beta, interleukin-4, or interleukin-10 suppressed in a dose-dependent manner the priming effects of tumor necrosis factor-alpha and interferon-gamma. The results of this study have implications for understanding the mechanisms by which cytokines and microglia contribute to processes of host defense and neurodegeneration via generation of reactive oxygen intermediates.