Toward the Development of Standards for Yellow Flashing Lights Used in Work Zones.

Flashing yellow warning lights are important for worker and driver safety in work zones. Current standards for these lights do not address whether and how they should be coordinated to provide course-way information to drivers navigating through work zones. A field study in which the intensities and flash patterns of warning lights along a simulated work zone were varied during daytime and nighttime, was conducted to assess drivers' responses to different configurations, leading to several conclusions. During the daytime, driver responses were relatively insensitive to warning light characteristics, although they preferred sequential and synchronized flash patterns over random, uncoordinated flashing. At nighttime, a temporal peak intensity of 25 cd with a sequential flash pattern was optimal for providing course-way information. A single initial warning light having a higher intensity may help drivers detect the work zone without creating unacceptable visual discomfort.

Vancomycin-intermediate Staphylococcus aureus isolates are attenuated for virulence when compared with susceptible progenitors.

OBJECTIVES: Vancomycin-intermediate Staphylococcus aureus (VISA) is associated with genetic changes that may also impact upon pathogenicity. In the current study, we compared the virulence of clinical VISA strains with their isogenic vancomycin-susceptible progenitors (VSSA). METHODS: Production of the critical virulence protein, α toxin, was assessed using Western blot analysis and was correlated to agr activity using a bioluminescent agr-reporter. Cytotoxicity and intracellular persistence were compared ex vivo for VSSA and VISA within non-professional phagocytes (NPP). Virulence and host immune responses were further explored in vivo using a murine model of bacteraemia. RESULTS: VISA isolates produced up to 20-fold less α toxin compared with VSSA, and this was corroborated by either loss of agr activity due to agr mutation, or altered agr activity in the absence of mutation. VISA were less cytotoxic towards NPP and were associated with enhanced intracellular persistence, suggesting that NPP may act as a reservoir for VISA. Infection with VSSA strains produced higher mortality in a murine bacteraemia model (≥90% 7-day mortality) compared with infection with VISA isolates (20% to 50%, p <0.001). Mice infected with VISA produced a dampened immune response (4.6-fold reduction in interleukin-6, p <0.001) and persistent organ bacterial growth was observed for VISA strains out to 7 days. CONCLUSIONS: These findings highlight the remarkable adaptability of S. aureus, whereby, in addition to having reduced antibiotic susceptibility, VISA alter the expression of pathogenic factors to circumvent the host immune response to favour persistent infection over acute virulence.

Toll-IL1 receptor-mediated innate immune responses vary across HBV genotype and predict treatment response to pegylated-IFN in HBeAg-positive CHB patients.

Patients with hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) have suppressed TLR2 expression, function and cytokine production. The aim of this study was to explore the importance of hepatitis B virus (HBV) genotype in innate immune responses and investigate whether Toll-like receptor (TLR) expression/function has potential roles as predictive biomarkers of successful therapy with pegylated interferon (Peg-IFN) therapy of HBeAg seroconversion in HBeAg-positive patients. We showed that as early as 4 weeks after initiation of Peg-IFN, future HBeAg seroconverters had significantly elevated levels of TLR2 expression on monocytes. TLR2-associated IL-6 production at baseline and week 4 of therapy and TLR4 IL-6 production at week 4 were also markedly elevated in HBeAg seroconverters. HBV genotype also influenced treatment response, with genotypes A and B more likely to seroconvert than D. We were able to demonstrate that these differences were due in part to the interaction of the specific HBeAg proteins with TLR pathway adaptor molecules, and these interactions were genotype dependent. HBeAg-mediated modulation of TLR signalling was also observed in Huh7 cells, following stimulation with Pam3Cys. Importantly, the addition of IFN-α to TLR2-stimulated cells cotransfected with an HBeAg expression plasmid reversed HBeAg-mediated suppression of hepatocytes. These findings demonstrate that patients with an activated inflammatory response are much more likely to respond to IFN therapy, with TLR responses showing promise as potential biomarkers of HBeAg seroconversion in this setting. Furthermore, our findings suggest there is differential genotype-specific HBeAg suppression of innate signalling pathways which may account for some of the clinical differences observed across the CHB spectrum.

Downregulation of interleukin-18-mediated cell signaling and interferon gamma expression by the hepatitis B virus e antigen.

UNLABELLED: The mechanisms by which hepatitis B virus (HBV) establishes and maintains chronic hepatitis B infection (CHB) are poorly defined. Innate immune responses play an important role in reducing HBV replication and pathogenesis. HBV has developed numerous mechanisms to escape these responses, including the production of the secreted hepatitis B e antigen (HBeAg), which has been shown to regulate antiviral toll-like receptor (TLR) and interleukin-1 (IL-1) signaling. IL-18 is a related cytokine that inhibits HBV replication in hepatoma cell lines and in the liver through the induction of gamma interferon (IFN-γ) by NK cells and T cells. We hypothesized that HBV or HBV proteins inhibit IFN-γ expression by NK cells as an accessory immunomodulatory function. We show that HBeAg protein inhibits the NF-κB pathway and thereby downregulates NK cell IFN-γ expression. Additionally, IFN-γ expression was significantly inhibited by exposure to serum from individuals with HBeAg-positive but not HBeAg-negative chronic HBV infection. Further, we show that the HBeAg protein suppresses IL-18-mediated NF-κB signaling in NK and hepatoma cells via modulation of the NF-κB pathway. Together, these findings show that the HBeAg inhibits IL-18 signaling and IFN-γ expression, which may play an important role in the establishment and/or maintenance of persistent HBV infection. IMPORTANCE: It is becoming increasingly apparent that NK cells play a role in the establishment and/or maintenance of chronic hepatitis B infection. The secreted HBeAg is an important regulator of innate and adaptive immune responses. We now show that the HBeAg downregulates NK cell-mediated IFN-γ production and IL-18 signaling, which may contribute to the establishment of infection and/or viral persistence. Our findings build on previous studies showing that the HBeAg also suppresses the TLR and IL-1 signaling pathways, suggesting that this viral protein is a key regulator of antiviral innate immune responses.

Toll-like receptor 3 and 7/8 function is impaired in hepatitis C rapid fibrosis progression post-liver transplantation.

Recurrence of hepatitis C (HCV) postliver transplant is universal, with a subgroup developing rapid hepatic fibrosis. Toll-like receptors (TLRs) are critical to innate antiviral responses and HCV alters TLR function to evade immune clearance. Whether TLRs play a role in rapid HCV recurrence posttransplant is unknown. We stimulated peripheral blood mononuclear cells (PBMCs) from 70 patients with HCV postliver transplant with TLR subclass-specific ligands and measured cytokine production, TLR expression and NK cell function. Rate of fibrosis progression was calculated using posttransplant liver biopsies graded by Metavir scoring (F0-4; R=fibrosis stage/year posttransplant; rapid fibrosis defined as >0.4 units/year). Thirty of 70 (43%) patients had rapid fibrosis progression. PBMCs from HCV rapid-fibrosers produced less IFNα with TLR7/8 stimulation (p=0.039), less IL-6 at baseline (p=0.027) and with TLR3 stimulation (p=0.008) and had lower TLR3-mediated monocyte IL-6 production (p=0.028) compared with HCV slow fibrosers. TLR7/8-mediated NKCD56 dim cell secretion of IFNγ was impaired in HCV rapid fibrosis (p=0.006) independently of IFNα secretion and TLR7/8 expression, while cytotoxicity remained preserved. Impaired TLR3 and TLR7/8-mediated cytokine responses may contribute to aggressive HCV recurrence postliver transplantation through impaired immune control of HCV and subsequent activation of fibrogenesis.

Inter- and intra-laboratory study to determine the reproducibility of toxicogenomics datasets.

The application of toxicogenomics as a predictive tool for chemical risk assessment has been under evaluation by the toxicology community for more than a decade. However, it predominately remains a tool for investigative research rather than for regulatory risk assessment. In this study, we assessed whether the current generation of microarray technology in combination with an in vitro experimental design was capable of generating robust, reproducible data of sufficient quality to show promise as a tool for regulatory risk assessment. To this end, we designed a prospective collaborative study to determine the level of inter- and intra-laboratory reproducibility between three independent laboratories. All test centres (TCs) adopted the same protocols for all aspects of the toxicogenomic experiment including cell culture, chemical exposure, RNA extraction, microarray data generation and analysis. As a case study, the genotoxic carcinogen benzo[a]pyrene (B[a]P) and the human hepatoma cell line HepG2 were used to generate three comparable toxicogenomic data sets. High levels of technical reproducibility were demonstrated using a widely employed gene expression microarray platform. While differences at the global transcriptome level were observed between the TCs, a common subset of B[a]P responsive genes (n=400 gene probes) was identified at all TCs which included many genes previously reported in the literature as B[a]P responsive. These data show promise that the current generation of microarray technology, in combination with a standard in vitro experimental design, can produce robust data that can be generated reproducibly in independent laboratories. Future work will need to determine whether such reproducible in vitro model(s) can be predictive for a range of toxic chemicals with different mechanisms of action and thus be considered as part of future testing regimes for regulatory risk assessment.

Hepatic TLR2 & TLR4 expression correlates with hepatic inflammation and TNF-α in HCV & HCV/HIV infection.

Signalling activated by Toll-like receptors (TLRs) can result in the production of tumour necrosis factor alpha (TNF-α) which is implicated in hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infection. No study has examined or compared hepatic expression of TLRs in both HCV and HCV/HIV. Liver and peripheral blood mononuclear cells (PBMCs) were obtained from HCV & HCV/HIV-infected patients and PBMCs from HIV-infected patients. Liver RNA was analysed by microarray and reverse transcription quantitative PCR (RT-qPCR). PBMCs were analysed by flow cytometry. Associations with hepatic histology and infection type were sought. Forty-six HCV, 20 HIV and 27 HCV/HIV-infected patients were recruited. Increasing Metavir inflammatory activity score was associated with increased hepatic TLR mRNA by RT-qPCR: TLR2 (P ≤ 0.001), TLR4 (P = 0.008) and TNF-α (P ≤ 0.001). A high degree of correlation was seen between hepatic mRNA expression of TNF-αvs TLR2 (r(2) = 0.66, P < 0.0001) and TLR4 (r(2) = 0.60, P < 0.0001). No differences in TLR gene or protein expression was observed between HCV, HCV/HIV- or HIV-infected groups. Hepatic TLR2, TLR4 and TNF-α mRNA are associated with hepatic inflammation in both HCV and HCV/HIV infection. High correlation between TNF-α and TLR2/TLR4 suggests a role for the innate immune response in TNF-α production. Activation of the innate immune response appears to be independent of infection type.

Toll-like receptor expression in chronic hepatitis C: correlation with pro-inflammatory cytokine levels and liver injury.

BACKGROUND/AIMS: Toll-like receptors (TLR's) are critical receptors that promote innate immune responses to pathogen-associated molecular patterns. Activation of TLR's leads to production of pro-inflammatory cytokines such as tumour necrosis factor (TNF)-alpha. This study investigates whether peripheral blood monocyte expression of TLR's is disturbed in patients with chronic hepatitis C and whether levels of expression of these molecules are significantly correlated with hepatitis C virus (HCV) genotype, viral load, hepatic necroinflammatory activity, histological stage and circulating TNF-alpha concentrations. METHODS: In 18 non-cirrhotic patients with biopsy-proven, virologically-confirmed chronic hepatitis C and 32 controls, we measured expression of TLR2 and TLR4 on peripheral blood monocytes. HCV genotype, viral load, serum alanine aminotransferase (ALT) levels, histological stage of disease and circulating TNF-alpha and endotoxin levels were also determined. RESULTS: Peripheral blood monocyte expression of TLR2 and TLR4 were significantly increased in patients with chronic hepatitis C compared to controls, irrespective of HCV genotype or histological stage of disease. Circulating levels of TNF-alpha were also significantly increased in patients with chronic hepatitis C. In both the overall study cohort and patients with chronic hepatitis C, monocyte expression of TLR2, but not of TLR4, correlated significantly with serum TNF-alpha levels. In patients with chronic hepatitis C, monocyte expression of TLR2, but not of TLR4, also correlated significantly with serum ALT levels. Expression of TLR's was not significantly correlated with viral load. CONCLUSIONS: Up-regulation of peripheral blood monocyte expression of TLR2 and TLR4 occurs in patients with chronic hepatitis C. Increased monocyte expression of TLR2, but not of TLR4, correlates significantly with both increased circulating TNF-alpha levels and hepatic necroinflammatory activity in this disorder.

Regulation of Toll-like receptor (TLR)2 and TLR4 on CD14dimCD16+ monocytes in response to sepsis-related antigens.

Rapid overproduction of proinflammatory cytokines are characteristic of sepsis. CD14(dim)CD16(+) monocytes are thought to be major producers of cytokine and have been shown to be elevated in septic patients. Toll-like receptors (TLR) are pattern recognition receptors important in mediating the innate immune response and their activation can lead to production of cytokines. Using whole blood culture and flow cytometry we have investigated TLR2 and TLR4 regulation after stimulation with sepsis-relevant antigens [lipopolysaccharide (LPS), Staphylococcal enterotoxin B (SEB) and peptidoglycan (PGN)]. The percentage of CD14(dim)CD16(+) monocyte population expanded at 20 h post-stimulation, after a rise in tumour necrosis factor (TNF)-alpha and interleukin (IL)-6 at 2 h. A strong positive correlation between the percentage of CD14(dim)CD16(+) monocytes and secreted TNF-alpha was demonstrated (r = 0.72). Furthermore, we were able to induce expansion of the CD14(dim)CD16(+) population to approximately 35% of all monocytes with the addition of recombinant TNF-alpha to the whole blood culture. TLR4 was found to be expressed 2.5 times higher on CD14(dim)CD16(+) compared to CD14(+) CD16(-) monocytes, while TLR2 expression was similar in both subpopulations. The CD14(dim)CD16(+) and CD14(+) CD16(-) monocyte populations were different in their response to various antigens. LPS down-regulated TLR4 by 4.9 times in CD16(+) monocytes compared to only 2.3 times in CD16(-) monocytes at 2 h. LPS was able to up-regulate TLR2 by 6.2 times after 2 h, with no difference between the subpopulations. LPS further up-regulated TLR2 by 18.4 times after 20 h only in the CD14(+) CD16(-) population. PGN and SEB induced no significant changes in TLR2 or TLR4 expression. We hypothesize that following exposure to bacterial antigens, subsequent TNF-alpha drives a differentiation of monocytes into a CD14(dim)CD16(+) subpopulation.

Lymphoepithelioma-like carcinoma of the uterine cervix.

We describe two rare cervical tumors having morphologic features closely resembling those of the nasopharyngeal lymphoepithelioma. This entity has historically been classified as a subtype of squamous cell carcinoma, but after reviewing the literature and the two cases presented here, we propose that this tumor is a distinct carcinoma of the cervix that differs from squamous cell carcinoma in that it carries a more favorable prognosis, typically affects a younger population of women, is more prevalent in noncaucasian populations (especially those of Asian descent), and lacks a clearly defined association with infection due to human papilloma virus (HPV).

Analytical far-field divergence angle of a truncated gaussian beam.

Approximate, but accurate, analytical expressions for the far-field divergence angle of a Gaussian beam normally incident on a circular aperture are derived. A first equation is obtained based on the concept of Gaussian transform, in which the Bessel function present in the far-field diffraction integral is approximated by a Gaussian function. Refining this approach yields another simple, practical closed-form formula with such a level of accuracy that we propose that it can be used as an exact reference. All approximations hold for any combination of Gaussian beam width and aperture radius.

Temporal characteristics of evaluation anxiety.

The temporal characteristics of evaluation anxiety are not well-defined by previous research. We examined the effects of length of the pre-evaluation interval (3, 6, or 12 minutes) and stage of the pre-evaluation interval at which evaluation anxiety was measured (start, middle, or end) on evaluation anxiety while participants performed an activity that was the focus of the impending evaluation. Participants wrote their opinion on a controversial social issue while anticipating a subject matter expert's judgment of their social maturity, and evaluation anxiety was measured by a battery of state anxiety measures. Higher levels of evaluation anxiety were detected on the Worry-Emotionality Questionnaire (WEQ) Worry subscale at the end of the pre-evaluation interval than at earlier stages, regardless of interval length, although individual difference variables exerted an important influence. Individuals with high trait self-presentation concerns experienced particularly high state anxiety (State-Trait Anxiety Inventory) at the start of the longest pre-evaluation interval. Low self-efficacy individuals showed a U-shaped pattern across pre-evaluation stages on both the WEQ Worry and Emotionality subscales, while high self-efficacy participants showed either no change (worry) or an inverted-U pattern (emotionality). Implications for the experimental measurement of evaluation anxiety were discussed.

Relationship between selected orientation rest frame, circular vection and space motion sickness.

Space motion sickness (SMS) and spatial orientation and motion perception disturbances occur in 70-80% of astronauts. People select "rest frames" to create the subjective sense of spatial orientation. In microgravity, the astronaut's rest frame may be based on visual scene polarity cues and on the internal head and body z axis (vertical body axis). The data reported here address the following question: Can an astronaut's orientation rest frame be related and described by other variables including circular vection response latencies and space motion sickness? The astronaut's microgravity spatial orientation rest frames were determined from inflight and postflight verbal reports. Circular vection responses were elicited by rotating a virtual room continuously at 35 degrees/s in pitch, roll and yaw with respect to the astronaut. Latency to the onset of vection was recorded from the time the crew member opened their eyes to the onset of vection. The astronauts who used visual cues exhibited significantly shorter vection latencies than those who used internal z axis cues. A negative binomial regression model was used to represent the observed total SMS symptom scores for each subject for each flight day. Orientation reference type had a significant effect, resulting in an estimated three-fold increase in the expected motion sickness score on flight day 1 for astronauts who used visual cues. The results demonstrate meaningful classification of astronauts' rest frames and their relationships to sensitivity to circular vection and SMS. Thus, it may be possible to use vection latencies to predict SMS severity and duration.

Self-motion perception: assessment by computer-generated animations.

The goal of this research is more precise description of adaptation to sensory rearrangements, including microgravity, by development of improved procedures for assessing spatial orientation perception. Thirty-six subjects reported perceived self-motion following exposure to complex inertial-visual motion. Twelve subjects were assigned to each of 3 perceptual reporting procedures: (a) animation movie selection, (b) written report selection and (c) verbal report generation. The question addressed was: do reports produced by these procedures differ with respect to complexity and reliability? Following repeated (within-day and across-day) exposures to 4 different "motion profiles," subjects either (a) selected movies presented on a laptop computer, or (b) selected written descriptions from a booklet, or (c) generated self-motion verbal descriptions that corresponded most closely with their motion experience. One "complexity" and 2 reliability "scores" were calculated. Contrary to expectations, reliability and complexity scores were essentially equivalent for the animation movie selection and written report selection procedures. Verbal report generation subjects exhibited less complexity than did subjects in the other conditions and their reports were often ambiguous. The results suggest that, when selecting from carefully written descriptions and following appropriate training, people may be better able to describe their self-motion experience with words than is usually believed.

Failure to support a test for penis envy.

Johnson (1966) reported that significantly more women than men did not comply with the request to return their special machine-scoring pencils after completing a final examination in introductory psychology. In the present replication, Johnson's conclusion that keeping the pencils may be manifestation of penis envy was not supported.

Hypochondria in women as a function of birth order.

The hypothesis that firstborn women would score higher than later-born women on a measure of hypochondria was supported. This result is explained in terms of parental treatment, specifically, the modeling of greater concern about the health of their girls by inexperienced parents.