Antioxidant-Induced Changes of the AP-1 Transcription Complex Are Paralleled by a Selective Suppression of Human Papillomavirus Transcription

Although E6 and E7 themselves possess intrinsic trans-activation capacity on their homologous promoters (14, 52), constitutive expression of E6 or E7 in immortalized or malignantly transformed human keratinocytes is mainly dependent on the availability of a defined set of transcription factors derived from the infected host cell. AP-1, for example, normally consisting of a heterodimer between c-Fos and c-Jun (for a review, see reference 4), seems to play a central role in transcriptional regulation of viral oncogene expression, since point mutations of the corresponding consensus sequences within the upstream regulatory region (URR) of HPV-16 or HPV-18 almost completely abolish the expression of URR-driven reporter plasmids in transient transfection assays Recent studies have shown that trans-activation and the DNA-binding affinity of AP-1 (1, 44) as well as that of other transcription factors such as NF-B (65, 78) or p53 (29) can be modulated not only via posttranslational modifications such as phosphorylation or dephosphorylation (for a review, see ref- erence 31) but also by alterations of the intracellular redox status. This can be achieved by certain cytokines, which are able to induce a prooxidant state within the cell by generating reactive oxygen intermediates (ROIs) Similar prooxidant conditions can also be generated either by exposing cells directly to hydrogen peroxide (40) or by genotoxic stress after UV irradiation (60), leading in both cases to the activation of AP-1 (74) and NF-B (44). While such intracellular redox changes are normally counterbalanced by antioxidant enzymes like Cu-Zn-superoxide dismutases (48) or oxidoreductases such as thioredoxin In a previous study, we showed that tumor necrosis factor alpha-induced transcription of a chemokine gene encoding monocyte chemoattractant protein 1 (MCP-1) could be completely abrogated when nonmalignant HPV-positive cells were incubated with PDTC prior to cytokine addition (57). Consistent with the aforementioned model of redox modulation of specific genes, detailed promoter analysis has revealed that NF-B and AP-1 are indeed the predominant transcription factors involved in the regulation of this particular chemokine Since AP-1 is also a central transcription factor for efficien

Quantum-Dense Metrology

Quantum metrology utilizes entanglement for improving the sensitivity of measurements. Up to now the focus has been on the measurement of just one out of two non-commuting observables. Here we demonstrate a laser interferometer that provides information about two non-commuting observables, with uncertainties below that of the meter's quantum ground state. Our experiment is a proof-of-principle of quantum dense metrology, and uses the additional information to distinguish between the actual phase signal and a parasitic signal due to scattered and frequency shifted photons. Our approach can be readily applied to improve squeezed-light enhanced gravitational-wave detectors at non-quantum noise limited detection frequencies in terms of a sub shot-noise veto-channel.Comment: 5 pages, 3 figures; includes supplementary material

Captive reptile mortality rates in the home and implications for the wildlife trade

The trade in wildlife and keeping of exotic pets is subject to varying levels of national and international regulation and is a topic often attracting controversy. Reptiles are popular exotic pets and comprise a substantial component of the live animal trade. High mortality of traded animals raises welfare concerns, and also has implications for conservation if collection from the wild is required to meet demand. Mortality of reptiles can occur at any stage of the trade chain from collector to consumer. However, there is limited information on mortality rates of reptiles across trade chains, particularly amongst final consumers in the home. We investigated mortality rates of reptiles amongst consumers using a specialised technique for asking sensitive questions, additive Randomised Response Technique (aRRT), as well as direct questioning (DQ). Overall, 3.6% of snakes, chelonians and lizards died within one year of acquisition. Boas and pythons had the lowest reported mortality rates of 1.9% and chameleons had the highest at 28.2%. More than 97% of snakes, 87% of lizards and 69% of chelonians acquired by respondents over five years were reported to be captive bred and results suggest that mortality rates may be lowest for captive bred individuals. Estimates of mortality from aRRT and DQ did not differ significantly which is in line with our findings that respondents did not find questions about reptile mortality to be sensitive. This research suggests that captive reptile mortality in the home is rather low, and identifies those taxa where further effort could be made to reduce mortality rate

The role of clathrin in post-golgi trafficking in toxoplasma gondii

Apicomplexan parasites are single eukaryotic cells with a highly polarised secretory system that contains unique secretory organelles (micronemes and rhoptries) that are required for host cell invasion. In contrast, the role of the endosomal system is poorly understood in these parasites. With many typical endocytic factors missing, we speculated that endocytosis depends exclusively on a clathrin-mediated mechanism. Intriguingly, in Toxoplasma gondii we were only able to observe the endogenous clathrin heavy chain 1 (CHC1) at the Golgi, but not at the parasite surface. For the functional characterisation of Toxoplasma gondii CHC1 we generated parasite mutants conditionally expressing the dominant negative clathrin Hub fragment and demonstrate that CHC1 is essential for vesicle formation at the trans-Golgi network. Consequently, the functional ablation of CHC1 results in Golgi aberrations, a block in the biogenesis of the unique secretory microneme and rhoptry organelles, and of the pellicle. However, we found no morphological evidence for clathrin mediating endocytosis in these parasites and speculate that they remodelled their vesicular trafficking system to adapt to an intracellular lifestyle

Analysis of the IDS Gene in 38 Patients with Hunter Syndrome: The c.879G>A (p.Gln293Gln) Synonymous Variation in a Female Create Exonic Splicing

BACKGROUND: Hunter syndrome (mucopolysaccharidosis type II, MPS II) is a rare disease inherited in an X-linked autosomal recessive pattern. It is the prevailing form of the mucopolysaccharidoses in China. Here we investigated mutations of IDS (iduronate 2-sulfatase) gene in 38 unrelated Chinese patients, one of which is a female. METHODS: Peripheral leucocytes were collected from the patients and the IDS gene was amplified to looking for the variations. For a female patient, the X chromosome status was analyzed by androgen receptor X-inactivation assay and the mutation impact on RNA level was further performed by reverse transcription polymerase chain reaction. RESULTS: We discovered that point mutations constituted the major form while mutations in codon p.R468 defined the largest number of patients in our cohort. Consistent with data from other ethnic groups, exons 9 and 3 had comparatively more mutations, while exon 2 had quite a few mutations unique to Chinese patients. Of the 30 different mutations identified, only 9 were novel: one was a premature termination mutation, i.e., c.196C>T (p.Gln66X); three were missense mutations, i.e., c.200T>C (p.Leu67Pro), c.215T>C (p.Leu72Pro), c.389C>T (p.Thr130Ile); one was a small deletion, i.e., c.1104_1122del19 (p.Ser369ArgfsX16); and one was a deletion that spanned both exons 8 and 9 deletion leading to gross structural changes in the IDS gene. In addition, a synonymous mutation c.879G>A (p.Gln293Gln) was identified in a female Hunter disease patient, which resulted in loss of the original splicing site, activated a cryptic splicing site upstream, leading to a 28 bp deletion and a premature termination at p. Tyr285GlufsX47. Together with concurrent skewed X-inactivation this was believed to facilitate the development of Hunter disease in this girl. CONCLUSIONS: In conclusion, the molecular analysis of IDS gene in Chinese patients confirmed the Hunter disease diagnosis and expanded the mutation and clinical spectrum of this devastating disorder

Tissue Compartment Analysis for Biomarker Discovery by Gene Expression Profiling

BACKGROUND:Although high throughput technologies for gene profiling are reliable tools, sample/tissue heterogeneity limits their outcomes when applied to identify molecular markers. Indeed, inter-sample differences in cell composition contribute to scatter the data, preventing detection of small but relevant changes in gene expression level. To date, attempts to circumvent this difficulty were based on isolation of the different cell structures constituting biological samples. As an alternate approach, we developed a tissue compartment analysis (TCA) method to assess the cell composition of tissue samples, and applied it to standardize data and to identify biomarkers. METHODOLOGY/PRINCIPAL FINDINGS:TCA is based on the comparison of mRNA expression levels of specific markers of the different constitutive structures in pure isolated structures, on the one hand, and in the whole sample on the other. TCA method was here developed with human kidney samples, as an example of highly heterogeneous organ. It was validated by comparison of the data with those obtained by histo-morphometry. TCA demonstrated the extreme variety of composition of kidney samples, with abundance of specific structures varying from 5 to 95% of the whole sample. TCA permitted to accurately standardize gene expression level amongst >100 kidney biopsies, and to identify otherwise imperceptible molecular disease markers. CONCLUSIONS/SIGNIFICANCE:Because TCA does not require specific preparation of sample, it can be applied to all existing tissue or cDNA libraries or to published data sets, inasmuch specific operational compartments markers are available. In human, where the small size of tissue samples collected in clinical practice accounts for high structural diversity, TCA is well suited for the identification of molecular markers of diseases, and the follow up of identified markers in single patients for diagnosis/prognosis and evaluation of therapy efficiency. In laboratory animals, TCA will interestingly be applied to central nervous system where tissue heterogeneity is a limiting factor

Acute effects of remote ischemic preconditioning on cutaneous microcirculation - a controlled prospective cohort study

<p>Abstract</p> <p>Background</p> <p>Therapeutic strategies aiming to reduce ischemia/reperfusion injury by conditioning tissue tolerance against ischemia appear attractive not only from a scientific perspective, but also in clinics. Although previous studies indicate that remote ischemic intermittent preconditioning (RIPC) is a systemic phenomenon, only a few studies have focused on the elucidation of its mechanisms of action especially in the clinical setting. Therefore, the aim of this study is to evaluate the acute microcirculatory effects of remote ischemic preconditioning on a distinct cutaneous location at the lower extremity which is typically used as a harvesting site for free flap reconstructive surgery in a human in-vivo setting.</p> <p>Methods</p> <p>Microcirculatory data of 27 healthy subjects (25 males, age 24 ± 4 years, BMI 23.3) were evaluated continuously at the anterolateral aspect of the left thigh during RIPC using combined Laser-Doppler and photospectrometry (Oxygen-to-see, Lea Medizintechnik, Germany). After baseline microcirculatory measurement, remote ischemia was induced using a tourniquet on the contralateral upper arm for three cycles of 5 min.</p> <p>Results</p> <p>After RIPC, tissue oxygen saturation and capillary blood flow increased up to 29% and 35% during the third reperfusion phase versus baseline measurement, respectively (both p = 0.001). Postcapillary venous filling pressure decreased statistically significant by 16% during second reperfusion phase (p = 0.028).</p> <p>Conclusion</p> <p>Remote intermittent ischemic preconditioning affects cutaneous tissue oxygen saturation, arterial capillary blood flow and postcapillary venous filling pressure at a remote cutaneous location of the lower extremity. To what extent remote preconditioning might ameliorate reperfusion injury in soft tissue trauma or free flap transplantation further clinical trials have to evaluate.</p> <p>Trial registration</p> <p>ClinicalTrials.gov: <a href="http://www.clinicaltrials.gov/ct2/show/NCT01235286">NCT01235286</a></p

Prevention of Diabetes in NOD Mice by Repeated Exposures to a Contact Allergen Inducing a Sub-Clinical Dermatitis

BACKGROUND: Type 1 diabetes is an autoimmune disease, while allergic contact dermatitis although immune mediated, is considered an exposure driven disease that develops due to epicutaneous contact with reactive low-molecular chemicals. The objective of the present study was to experimentally study the effect of contact allergens on the development of diabetes in NOD mice. As the link between contact allergy and diabetes is yet unexplained we also examined the effect of provocation with allergens on Natural Killer T (NKT) cells, since involvement of NKT cells could suggest an innate connection between the two diseases. METHOD: NOD mice 4 weeks of age were exposed, on the ears, to two allergens, p-phenylenediamine and 2,4-dinitrochlorobenzene respectively, to investigate the diabetes development. The mice were followed for a maximum of 32 weeks, and they were either repeatedly exposed to the allergens or only sensitized a week after arrival. The stimulation of NKT cells by the two allergens were additionally studied in C57BL/6 mice. The mice were sensitized and two weeks later provocated with the allergens. The mice were subsequently euthanized at different time points after the provocation. RESULTS: It was found that repeated application of p-phenylenediamine reduced the incidence of diabetes compared to application with water (47% vs. 93%, P = 0.004). Moreover it was shown that in C57BL/6 mice both allergens resulted in a slight increment in the quantity of NKT cells in the liver. Application of the allergens at the same time resulted in an increased number of NKT cells in the draining auricular lymph node, and the increase appeared to be somewhat allergen specific as the accumulation was stronger for p-phenylenediamine. CONCLUSION: The study showed that repeated topical application on the ears with a contact allergen could prevent the development of diabetes in NOD mice. The contact allergens gave a non-visible, sub-clinical dermatitis on the application site. The preventive effect on diabetes may be due to stimulation of peripheral NKT cells, as shown for provocation with p-phenylenediamine in the C57BL/6 mouse. This epicutaneous procedure may lead to new strategies in prevention of type 1 diabetes in humans

Cell Lineage and Regional Identity of Cultured Spinal Cord Neural Stem Cells and Comparison to Brain-Derived Neural Stem Cells

Neural stem cells (NSCs) can be isolated from different regions of the central nervous system. There has been controversy whether regional differences amongst stem and progenitor cells are cell intrinsic and whether these differences are maintained during expansion in culture. The identification of inherent regional differences has important implications for the use of these cells in neural repair. Here, we compared NSCs derived from the spinal cord and embryonic cortex. We found that while cultured cortical and spinal cord derived NSCs respond similarly to mitogens and are equally neuronogenic, they retain and maintain through multiple passages gene expression patterns indicative of the region from which they were isolated (e.g Emx2 and HoxD10). Further microarray analysis identified 229 genes that were differentially expressed between cortical and spinal cord derived neurospheres, including many Hox genes, Nuclear receptors, Irx3, Pace4, Lhx2, Emx2 and Ntrk2. NSCs in the cortex express LeX. However, in the embryonic spinal cord there are two lineally related populations of NSCs: one that expresses LeX and one that does not. The LeX negative population contains few markers of regional identity but is able to generate LeX expressing NSCs that express markers of regional identity. LeX positive cells do not give rise to LeX-negative NSCs. These results demonstrate that while both embryonic cortical and spinal cord NSCs have similar self-renewal properties and multipotency, they retain aspects of regional identity, even when passaged long-term in vitro. Furthermore, there is a population of a LeX negative NSC that is present in neurospheres derived from the embryonic spinal cord but not the cortex

Profiles of Multidrug Resistance Protein-1 in the Peripheral Blood Mononuclear Cells of Patients with Refractory Epilepsy

BACKGROUND: About one third of patients with epilepsy become refractory to therapy despite receiving adequate medical treatment, possibly from multidrug resistance. P-glycoprotein, encoded by multidrug resistance protein-1 (MDR1) gene, at the blood brain barrier is considered as a major factor mediating drug efflux and contributing to resistance. Given that peripheral blood mononuclear cells (PBMNCs) express MDR1, we investigated a MDR1 status of PBMNCs in various subsets of epilepsy patients and demonstrated their association with clinical characteristics. METHODOLOGY/PRINCIPAL FINDINGS: Clinical and MDR1 data were collected from 140 patients with epilepsy, 30 healthy volunteers, and 20 control patients taking anti-epileptic drugs. PBMNCs were isolated, and basal MDR1 levels and MDR1 conformational change levels were measured by flow cytometry. MDR1 profiles were analyzed according to various clinical parameters, including seizure frequency and number of medications used in epilepsy patients. Epilepsy patients had a higher basal MDR1 level than non-epilepsy groups (p<0.01). Among epilepsy patients, there is a tendency for higher seizure frequency group to have higher basal MDR1 level (p = 0.059). The MDR1 conformational change level was significantly higher in the high-medication-use group than the low-use group (p = 0.028). Basal MDR1 (OR = 1.16 [95% CI: 1.060-1.268]) and conformational change level (OR = 1.11 [95% CI: 1.02-1.20]) were independent predictors for seizure frequency and number of medications, respectively. CONCLUSIONS/SIGNIFICANCE: The MDR1 profile of PBMNCs is associated with seizure frequency and medication conditions in patients with epilepsy