Stable transduction of bovine TLR4 and bovine MD-2 into LPS-nonresponsive cells and soluble CD14 promote the ability to respond to LPS

The interaction of bovine cells with lipopolysaccharide (LPS) was explored using human embryo kidney (HEK) 293 cell line stably transduced with bovine toll-like receptor-4 (TLR4) alone or in combination with bovine MD-2. These lines and mock-transduced HEK293 cells were tested by flow cytometry for LPS-fluorescein isothiocyanate (LPS-FITC) binding, nuclear factor kappa B (NFkappaB) activation, interleukin-8 (IL-8) production and interferon-beta mRNA expression/interferon (IFN) type I production. Whereas bovine TLR4 was sufficient to promote binding of high concentrations of LPS-FITC, both bovine TLR4 and MD-2 were required for activation by LPS, as assessed by NFkappaB activation and IL-8 production. Induction of IFN bioactivity was not observed in doubly transduced HEK293 cells, and no evidence for IFN-beta mRNA induction in response to LPS was obtained, although cells responded by IFN-beta mRNA expression to stimulation by Sendai virus and poly-inosinic acid-poly-cytidylic acid (poly(I:C)). Cells stably transduced with both bovine TLR4 and bovine MD-2 responded to LPS by IL-8 production, in decreasing order, in the presence of fetal bovine serum (FCS), of human serum, and of human serum albumin (HSA). The reduced activity in the presence of HSA could be restored by the addition of soluble CD14 (sCD14) but not of LPS binding protein (LBP). This is in contrast to macrophages which show a superior response to LPS in the presence of HSA when compared with macrophages stimulated by LPS in the presence of FCS. This suggests that macrophages but not HEK293 cells express factors rendering LPS stimulation serum-independent. Stably double-transduced cells reacted, in decreasing order, to LPS from Rhodobacter sphaeroides, to LPS from Escherichia coli, to synthetic lipd-IVa (compound 406), to diphosphoryl-lipid-A (S. minnesota) and to monophosphoryl-lipid-A (S. minnesota). They failed to react to the murine MD-2/TLR4 ligand taxol. This resembles the reactivity of bovine macrophages with regard to sensitivity (ED(50)) and order of potency but is distinct from the reactivity pattern of other species. This formally establishes that in order to react to LPS, cattle cells require serum factors (e.g. sCD14) and cell-expressed factors such as MD-2 and TLR4. The cell lines described are the first of a series expressing defined pattern recognition receptors (PRR) of bovine origin. They will be useful in the study of the interaction of the bovine TLR4-MD-2 complex and Gram-negative bovine pathogens, e.g. the agents causing Gram-negative bovine mastitis.

Comparison between confocal scanning laser tomography, scanning laser polarimetry and optical coherence tomography on the ability to detect localised retinal nerve fibre layer defects in glaucoma patients

BACKGROUND/AIM: To compare the ability of confocal scanning laser tomography (CSLT), scanning laser polarimetry (SLP) and optical coherence tomography (OCT) in recognising localised retinal nerve fibre layer (RNFL) defects. METHODS: 51 eyes from 43 patients with glaucoma were identified by two observers as having RNFL defects visible on optic disc photographs. 51 eyes of 32 normal subjects were used as controls. Three masked observers evaluated CSLT, SLP and OCT images to determine subjectively the presence of localised RNFL defects. RESULTS: Interobserver agreement was highest with OCT, followed by SLP and CSLT (mean kappa: 0.83, 0.69 and 0.64, respectively). RNFL defects were identified in 58.8% of CSLT, 66.7% of SLP and 54.9% of OCT (p = 0.02 between SLP and OCT) by at least two observers. In the controls, 94.1% of CSLT, 84.3% of SLP and 94.1% of OCT scans, respectively, were rated as normal (p = 0.02 between CSLT and SLP, and SLP and OCT). CONCLUSION: Approximately 20-40% of localised RNFL defects identified by colour optic disc photographs are not detected by CSLT, SPL or OCT. SLP showed a higher number of false-positive results than the other techniques, but also had a higher proportion of correctly identified RNFL defects in the glaucoma population.

Opinions of dating agents about strabismic subjects' ability to find a partner

AIMS: To determine the influence of strabismus on the ability to find a partner. METHODS: We interviewed Swiss dating agents retrieved from two Swiss online telephone directories using a validated questionnaire to determine whether strabismus has any impact on the ability to find a partner. During the interviews, subjects with internet access could view downloadable, digitally altered photographs of a strabismic man and women, as well as images of other computer-generated facial anomalies. RESULTS: Of the 40 dating agents, 92.5% judged that strabismic subjects have more difficulty finding a partner (p<0.001). Such difficulty was not associated with either gender or age but was perceived as being greater in exotropic than in esotropic persons (p<0.001). Among the seven facial disfigurements, strabismus was believed to have the third largest negative impact on finding a partner, after strong acne and a visible missing tooth. Dating agents also believed that potential partners perceive persons with strabismus as significantly less attractive (p<0.001), erotic (p<0.001), likeable (p<0.001), interesting (p<0.001), successful (p<0.001), intelligent (p = 0.001) and sporty (p = 0.01). CONCLUSIONS: Visible strabismus negatively influences the ability to find a partner. Because strabismus surgery in adults restores a normal functioning condition and reduces not only physical but also psychosocial difficulties, it cannot be considered a cosmetic procedure.

Multimodal approach in the use of clinical scoring, morphological MRI and biochemical T2-mapping and diffusion-weighted imaging in their ability to assess differences between cartilage repair tissue after microfracture therapy and matrix-associated autologous chondrocyte transplantation: a pilot study

OBJECTIVE: The aim of the present pilot study is to show initial results of a multimodal approach using clinical scoring, morphological magnetic resonance imaging (MRI) and biochemical T2-relaxation and diffusion-weighted imaging (DWI) in their ability to assess differences between cartilage repair tissue after microfracture therapy (MFX) and matrix-associated autologous chondrocyte transplantation (MACT). METHOD: Twenty patients were cross-sectionally evaluated at different post-operative intervals from 12 to 63 months after MFX and 12-59 months after MACT. The two groups were matched by age (MFX: 36.0+/-10.4 years; MACT: 35.1+/-7.7 years) and post-operative interval (MFX: 32.6+/-16.7 months; MACT: 31.7+/-18.3 months). After clinical evaluation using the Lysholm score, 3T-MRI was performed obtaining the MR observation of cartilage repair tissue (MOCART) score as well as T2-mapping and DWI for multi-parametric MRI. Quantitative T2-relaxation was achieved using a multi-echo spin-echo sequence; semi-quantitative diffusion-quotient (signal intensity without diffusion-weighting divided by signal intensity with diffusion weighting) was prepared by a partially balanced, steady-state gradient-echo pulse sequence. RESULTS: No differences in Lysholm (P=0.420) or MOCART (P=0.209) score were observed between MFX and MACT. T2-mapping showed lower T2 values after MFX compared to MACT (P=0.039). DWI distinguished between healthy cartilage and cartilage repair tissue in both procedures (MFX: P=0.001; MACT: P=0.007). Correlations were found between the Lysholm and the MOCART score (Pearson: 0.484; P=0.031), between the Lysholm score and DWI (Pearson:-0.557; P=0.011) and a trend between the Lysholm score and T2 (Person: 0.304; P=0.193). CONCLUSION: Using T2-mapping and DWI, additional information could be gained compared to clinical scoring or morphological MRI. In combination clinical, MR-morphological and MR-biochemical parameters can be seen as a promising multimodal tool in the follow-up of cartilage repair.

Disrupting the prefrontal cortex diminishes the human ability to build a good reputation.

Reputation formation pervades human social life. In fact, many people go to great lengths to acquire a good reputation, even though building a good reputation is costly in many cases. Little is known about the neural underpinnings of this important social mechanism, however. In the present study, we show that disruption of the right, but not the left, lateral prefrontal cortex (PFC) with low-frequency repetitive transcranial magnetic stimulation (rTMS) diminishes subjects' ability to build a favorable reputation. This effect occurs even though subjects' ability to behave altruistically in the absence of reputation incentives remains intact, and even though they are still able to recognize both the fairness standards necessary for acquiring and the future benefits of a good reputation. Thus, subjects with a disrupted right lateral PFC no longer seem to be able to resist the temptation to defect, even though they know that this has detrimental effects on their future reputation. This suggests an important dissociation between the knowledge about one's own best interests and the ability to act accordingly in social contexts. These results link findings on the neural underpinnings of self-control and temptation with the study of human social behavior, and they may help explain why reputation formation remains less prominent in most other species with less developed prefrontal cortices.

Ability to Work and Employment Rates in Human Immunodeficiency Virus (HIV)-1-Infected Individuals Receiving Combination Antiretroviral Therapy: The Swiss HIV Cohort Study

Background.  Limited data exist on human immunodeficiency virus (HIV)-infected individuals' ability to work after receiving combination antiretroviral therapy (cART). We aimed to investigate predictors of regaining full ability to work at 1 year after starting cART. Methods.  Antiretroviral-naive HIV-infected individuals <60 years who started cART from January 1998 through December 2012 within the framework of the Swiss HIV Cohort Study were analyzed. Inability to work was defined as a medical judgment of the patient's ability to work as 0%. Results.  Of 5800 subjects, 4382 (75.6%) were fully able to work, 471 (8.1%) able to work part time, and 947 (16.3%) were unable to work at baseline. Of the 947 patients unable to work, 439 (46.3%) were able to work either full time or part time at 1 year of treatment. Predictors of recovering full ability to work were non-white ethnicity (odds ratio [OR], 2.06; 95% confidence interval [CI], 1.20-3.54), higher education (OR, 4.03; 95% CI, 2.47-7.48), and achieving HIV-ribonucleic acid <50 copies/mL (OR, 1.83; 95% CI, 1.20-2.80). Older age (OR, 0.55; 95% CI, .42-.72, per 10 years older) and psychiatric disorders (OR, 0.24; 95% CI, .13-.47) were associated with lower odds of ability to work. Recovering full ability to work at 1 year increased from 24.0% in 1998-2001 to 41.2% in 2009-2012, but the employment rates did not increase. Conclusions.  Regaining full ability to work depends primarily on achieving viral suppression, absence of psychiatric comorbidity, and favorable psychosocial factors. The discrepancy between patients' ability to work and employment rates indicates barriers to reintegration of persons infected with HIV.

Opinion of headhunters about the ability of strabismic subjects to obtain employment

BACKGROUND: Through the opinion of Swiss headhunters, we wanted to determine the influence of strabismus on the ability to obtain employment. METHODS: Out of 31 randomly selected Swiss headhunters, 20 could be interviewed using a validated questionnaire. RESULTS: Forty-seven percent of the headhunters judged that strabismic subjects have more difficulties in obtaining a job. Gender has no influence on discrimination (p > 0.1). Asked about six facial disfigurements, strabismus was found to have the second largest negative impact on employment directly after acne. Strabismus was estimated to decrease the attractiveness of job applicants (p < 0.0001) and to have a negative impact on the overall judgment of a potential employer (p < 0.05). CONCLUSIONS: Visible strabismus influences negatively the ability to obtain a job. Because of its impact on the employability of a person, we believe that strabismus surgery in adults cannot be considered to be only a beautifying procedure.

Pericellular fibronectin is required for RhoA-dependent responses to cyclic strain in fibroblasts

To test the hypothesis that the pericellular fibronectin matrix is involved in mechanotransduction, we compared the response of normal and fibronectin-deficient mouse fibroblasts to cyclic substrate strain. Normal fibroblasts seeded on vitronectin in fibronectin-depleted medium deposited their own fibronectin matrix. In cultures exposed to cyclic strain, RhoA was activated, actin-stress fibers became more prominent, MAL/MKL1 shuttled to the nucleus, and mRNA encoding tenascin-C was induced. By contrast, these RhoA-dependent responses to cyclic strain were suppressed in fibronectin knockdown or knockout fibroblasts grown under identical conditions. On vitronectin substrate, fibronectin-deficient cells lacked fibrillar adhesions containing alpha5 integrin. However, when fibronectin-deficient fibroblasts were plated on exogenous fibronectin, their defects in adhesions and mechanotransduction were restored. Studies with fragments indicated that both the RGD-synergy site and the adjacent heparin-binding region of fibronectin were required for full activity in mechanotransduction, but not its ability to self-assemble. In contrast to RhoA-mediated responses, activation of Erk1/2 and PKB/Akt by cyclic strain was not affected in fibronectin-deficient cells. Our results indicate that pericellular fibronectin secreted by normal fibroblasts is a necessary component of the strain-sensing machinery. Supporting this hypothesis, induction of cellular tenascin-C by cyclic strain was suppressed by addition of exogenous tenascin-C, which interferes with fibronectin-mediated cell spreading.

Nanomechanics to drive stem cells in injured tissues: insights from current research and future perspectives

Stem cells reside within tissue, ensuring its natural ability to repair an injury. They are involved in the natural repair of damaged tissue, which encompasses a complex process requiring the modulation of cell survival, extracellular matrix turnover, angiogenesis, and reverse remodeling. To date, the real reparative potential of each tissue is underestimated and noncommittal. The assessment of the biophysical properties of the extracellular environment is an innovative approach to better understand mechanisms underlying stem cell function, and consequently to develop safe and effective therapeutic strategies replacing the loss of tissue. Recent studies have focused on the role played by biomechanical signals that drive stem cell death, differentiation, and paracrinicity in a genetic and/or an epigenetic manner. Mechanical stimuli acting on the shape can influence the biochemistry and gene expression of resident stem cells and, therefore, the magnitude of biological responses that promote the healing of injured tissue. Nanotechnologies have proven to be a revolutionary tool capable of dissecting the cellular mechanosensing apparatus, allowing the intercellular cross-talk to be decoded and enabling the reparative potential of tissue to be enhanced without manipulation of stem cells. This review highlights the most relevant findings of stem cell mechanobiology and presents a fascinating perspective in regenerative medicine.

Can MRI of the neck compete with clinical findings in assessing danger to life for survivors of manual strangulation? A statistical analysis

The aim of the study was to examine the clinical forensic findings of strangulation according to their ability to differentiate between life-threatening and non-life-threatening strangulation, compare clinical and MRI findings of the neck and discuss a simple score for life-threatening strangulation (SLS).

Role of meprins to protect ileal mucosa of Crohn's disease patients from colonization by adherent-invasive E. coli

Ileal lesions in Crohn's disease (CD) patients are colonized by pathogenic adherent-invasive Escherichia coli (AIEC) able to adhere to and invade intestinal epithelial cells (IEC), and to survive within macrophages. The interaction of AIEC with IEC depends on bacterial factors mainly type 1 pili, flagella, and outer membrane proteins. In humans, proteases can act as host defence mechanisms to counteract bacterial colonization. The protease meprin, composed of multimeric complexes of the two subunits alpha and beta, is abundantly expressed in IECs. Decreased levels of this protease correlate with the severity of the inflammation in patients with inflammatory bowel disease. The aim of the present study was to analyze the ability of meprin to modulate the interaction of AIEC with IECs. In patients with ileal CD we observed decreased levels of meprins, in particular that of meprin β. Dose-dependent inhibition of the abilities of AIEC strain LF82 to adhere to and invade intestinal epithelial T84 cells was observed when bacteria were pre-treated with both exogenous meprin α and meprin β. Dose-dependent proteolytic degradation of type 1 pili was observed in the presence of active meprins, but not with heat-inactivated meprins, and pretreatment of AIEC bacteria with meprins impaired their ability to bind mannosylated host receptors and led to decreased secretion of the pro-inflammatory cytokine IL-8 by infected T84 cells. Thus, decreased levels of protective meprins as observed in CD patients may contribute to increased AIEC colonization.

Spike-triggered reaction-time EEG as a possible assessment tool for driving ability

The impact of interictal epileptic activity (IEA) on driving is a rarely investigated issue. We analyzed the impact of IEA on reaction time in a pilot study. Reactions to simple visual stimuli (light flash) in the Flash test or complex visual stimuli (obstacle on a road) in a modified car driving computer game, the Steer Clear, were measured during IEA bursts and unremarkable electroencephalography (EEG) periods. Individual epilepsy patients showed slower reaction times (RTs) during generalized IEA compared to RTs during unremarkable EEG periods. RT differences were approximately 300 ms (p < 0.001) in the Flash test and approximately 200 ms (p < 0.001) in the Steer Clear. Prior work suggested that RT differences >100 ms may become clinically relevant. This occurred in 40% of patients in the Flash test and in up to 50% in the Steer Clear. When RT were pooled, mean RT differences were 157 ms in the Flash test (p < 0.0001) and 116 ms in the Steer Clear (p < 0.0001). Generalized IEA of short duration seems to impair brain function, that is, the ability to react. The reaction-time EEG could be used routinely to assess driving ability.