Differential effect of long versus short wavelength light exposure on pupillary re-dilation in patients with outer retinal disease.

BACKGROUND: In patients with outer retinal degeneration, a differential pupil response to long wavelength (red) versus short wavelength (blue) light stimulation has been previously observed. The goal of this study was to quantify differences in the pupillary re-dilation following exposure to red versus blue light in patients with outer retinal disease and compare them with patients with optic neuropathy and with healthy subjects. DESIGN: Prospective comparative cohort study. PARTICIPANTS: Twenty-three patients with outer retinal disease, 13 patients with optic neuropathy and 14 normal subjects. METHODS: Subjects were tested using continuous red and blue light stimulation at three intensities (1, 10 and 100 cd/m2) for 13 s per intensity. Pupillary re-dilation dynamics following the brightest intensity was analysed and compared between the three groups. MAIN OUTCOME MEASURES: The parameters of pupil re-dilation used in this study were: time to recover 90% of baseline size; mean pupil size at early and late phases of re-dilation; and differential re-dilation time for blue versus red light. RESULTS: Patients with outer retinal disease showed a pupil that tended to stay smaller after light termination and thus had a longer time to recovery. The differential re-dilation time was significantly greater in patients with outer retinal disease (median = 28.0 s, P < 0.0001) compared with controls and patients with optic neuropathy. CONCLUSIONS: A differential response of pupil re-dilation following red versus blue light stimulation is present in patients with outer retinal disease but is not found in normal eyes or among patients with visual loss from optic neuropathy.

Simultaneous cell morphometry and refractive index measurement with dual-wavelength digital holographic microscopy and dye-enhanced dispersion of perfusion medium.

Digital holographic microscopy (DHM) allows optical-path-difference (OPD) measurements with nanometric accuracy. OPD induced by transparent cells depends on both the refractive index (RI) of cells and their morphology. This Letter presents a dual-wavelength DHM that allows us to separately measure both the RI and the cellular thickness by exploiting an enhanced dispersion of the perfusion medium achieved by the utilization of an extracellular dye. The two wavelengths are chosen in the vicinity of the absorption peak of the dye, where the absorption is accompanied by a significant variation of the RI as a function of the wavelength.

Real-time dual-wavelength digital holographic microscopy for extended measurement range with enhanced axial resolution

We report on advanced dual-wavelength digital holographic microscopy (DHM) methods, enabling single-acquisition real-time micron-range measurements while maintaining single-wavelength interferometric resolution in the nanometer regime. In top of the unique real-time capability of our technique, it is shown that axial resolution can be further increased compared to single-wavelength operation thanks to the uncorrelated nature of both recorded wavefronts. It is experimentally demonstrated that DHM topographic investigation within 3 decades measurement range can be achieved with our arrangement, opening new applications possibilities for this interferometric technique. ©2008 COPYRIGHT SPIE

Submicrometer tomography of cells by multiple-wavelength digital holographic microscopy in reflection

We present first results on a method enabling mechanical scanning-free tomography with submicrometer axial resolution by multiple-wavelength digital holographic microscopy. By sequentially acquiring reflection holograms and summing 20 wavefronts equally spaced in spatial frequency in the 485-670 nm range, we are able to achieve a slice-by-slice tomographic reconstruction with a 0.6-1 mum axial resolution in a biological medium. The method is applied to erythrocytes investigation to retrieve the cellular membrane profile in three dimensions.

Tunable reporter signal production in feedback-uncoupled arsenic bioreporters.

Escherichia coli-based bioreporters for arsenic detection are typically based on the natural feedback loop that controls ars operon transcription. Feedback loops are known to show a wide range linear response to the detriment of the overall amplification of the incoming signal. While being a favourable feature in controlling arsenic detoxification for the cell, a feedback loop is not necessarily the most optimal for obtaining highest sensitivity and response in a designed cellular reporter for arsenic detection. Here we systematically explore the effects of uncoupling the topology of arsenic sensing circuitry on the developed reporter signal as a function of arsenite concentration input. A model was developed to describe relative ArsR and GFP levels in feedback and uncoupled circuitry, which was used to explore new ArsR-based synthetic circuits. The expression of arsR was then placed under the control of a series of constitutive promoters, which differed in promoter strength, and which could be further modulated by TetR repression. Expression of the reporter gene was maintained under the ArsR-controlled Pars promoter. ArsR expression in the systems was measured by using ArsR-mCherry fusion proteins. We find that stronger constitutive ArsR production decreases arsenite-dependent EGFP output from Pars and vice versa. This leads to a tunable series of arsenite-dependent EGFP outputs in a variety of systematically characterized circuitries. The higher expression levels and sensitivities of the response curves in the uncoupled circuits may be useful for improving field-test assays using arsenic bioreporters.

Dual-wavelength digital holographic microscopy with sub-nanometer axial accuracy

We present dual-wavelength Digital Holographic Microscopy (DHM) measurements on a certified 8.9 nm high Chromium thin step sample and demonstrate sub-nanometer axial accuracy. We introduce a modified DHM Reference Calibrated Hologram (RCH) reconstruction algorithm taking into account amplitude contributions. By combining this with a temporal averaging procedure and a specific dual-wavelength DHM arrangement, it is shown that specimen topography can be measured with an accuracy, defined as the axial standard deviation, reduced to at least 0.9 nm. Indeed, it is reported that averaging each of the two wavefronts recorded with real-time dual-wavelength DHM can provide up to 30% spatial noise reduction for the given configuration, thanks to their non-correlated nature. ©2008 COPYRIGHT SPIE

The Discrimination of Colored Acrylic, Cotton, and Wool Textile Fibers Using Micro-Raman Spectroscopy. Part 1 : In Situ Detection and Characterization of Dyes

Raman spectroscopy has been applied to characterize fiber dyes and determine the discriminating ability of the method. Black, blue, and red acrylic, cotton, and wool samples were analyzed. Four excitation sources were used to obtain complementary responses in the case of fluorescent samples. Fibers that did not provide informative spectra using a given laser were usually detected using another wavelength. For any colored acrylic, the 633-nm laser did not provide Raman information. The 514-nm laser provided the highest discrimination for blue and black cotton, but half of the blue cottons produced noninformative spectra. The 830-nm laser exhibited the highest discrimination for red cotton. Both visible lasers provided the highest discrimination for black and blue wool, and NIR lasers produced remarkable separation for red and black wool. This study shows that the discriminating ability of Raman spectroscopy depends on the fiber type, color, and the laser wavelength.

Mutations in NR2E3 can cause dominant or recessive retinal degenerations in the same family.

NR2E3, a photoreceptor-specific nuclear receptor (PNR), represses cone-specific genes and activates several rod-specific genes. In humans, mutations in NR2E3 have been associated with the recessively-inherited enhanced short-wavelength sensitive S-cone syndrome (ESCS) and, recently, with autosomal dominant (ad) retinitis pigmentosa (RP) (adRP). In the present work, we describe two additional families affected by adRP that carry a heterozygous c.166G&gt;A (p.G56R) mutation in the NR2E3 gene. Functional analysis determined the dominant negative activity of the p.G56R mutant protein as the molecular mechanism of adRP. Interestingly, in one pedigree, the most common causal variant for ESCS (p.R311Q) cosegregated with the adRP-linked p.G56R mutation, and the compound heterozygotes exhibited an ESCS-like phenotype, which in 1 of the 2 cases was strikingly "milder" than the patients carrying the p.G56R mutation alone. Impaired repression of cone-specific genes by the corepressors atrophin-1 (dentatorubral-pallidoluysian atrophy [DRPLA] gene product) and atrophin-2 (arginine-glutamic acid dipeptide repeat [RERE] protein) appeared to be a molecular mechanism mediating the beneficial effect of the p.R311Q mutation. Finally, the functional dominance of the p.R311Q variant to the p.G56R mutation is discussed.

In vivo measurement of tissue oxygenation by time-resolved luminescence spectroscopy: advantageous properties of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate.

Measuring tissue oxygenation in vivo is of interest in fundamental biological as well as medical applications. One minimally invasive approach to assess the oxygen partial pressure in tissue (pO2) is to measure the oxygen-dependent luminescence lifetime of molecular probes. The relation between tissue pO2 and the probes' luminescence lifetime is governed by the Stern-Volmer equation. Unfortunately, virtually all oxygen-sensitive probes based on this principle induce some degree of phototoxicity. For that reason, we studied the oxygen sensitivity and phototoxicity of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate [Ru(Phen)] using a dedicated optical fiber-based, time-resolved spectrometer in the chicken embryo chorioallantoic membrane. We demonstrated that, after intravenous injection, Ru(Phen)'s luminescence lifetime presents an easily detectable pO2 dependence at a low drug dose (1 mg∕kg) and low fluence (120 mJ∕cm2 at 470 nm). The phototoxic threshold was found to be at 10 J∕cm2 with the same wavelength and drug dose, i.e., about two orders of magnitude larger than the fluence necessary to perform a pO2 measurement. Finally, an illustrative application of this pO2 measurement approach in a hypoxic tumor environment is presented.

Transcriptional Activation and Receptor Dimerization is Affected by Mutations in the NR2E3 Ligand-Binding Domain

Purpose: Mutations in the ligand-binding domain (LBD) of NR2E3 cause recessively inherited enhanced short wavelength sensitive (S-) cone syndrome (ESCS), Goldmann-Favre syndrome (GFS) and clumped pigmentary retinal degeneration (CPRD). In addition to ligand binding, the LBD contains also essential amino acid sequences for the oligomerization of nuclear receptors. The aim of our studies is to characterize the impact of mutations in the LBD on receptor oligomerization and transcriptional activity of NR2E3. Methods: The different NR2E3 mutants were generated by QuickChange mutagenesis and analyzed in 293T-based transactivation studies and BRET2 (bioluminescence resonance electron transfer) assays. In silico homology modeling of mutant proteins was also performed using available crystallographic data of related nuclear receptors. Results: The mutants p.W234S, p.A256V, p.A256E, p.L263P, p.R309G, p.R311Q, p.R334G, p.L336P, p.L353V, p.R385P and p.M407K, all located in the LBD, showed impaired receptor dimerization at various degrees. Impaired repressor dimerization as assessed by BRET2 assays did not always correlate with impaired repressor function of NR2E3 as assessed by cell-based reporter assays. There were minor differences of transcriptional activity of mutant proteins on mouse S-opsin (opn1sw), mouse cone arrestin (arr3) and human cone arrestin, suggesting that the effect of LBD mutations was independent of the promoter context. Conclusions: Mutational analysis and homology modeling allowed the characterization of potential oligomerization interfaces of the NR2E3 LBD. Additionally, mutations in NR2E3 LBD may cause recessive retinal degenerations by different molecular mechanisms.

Chromatic pupil responses: preferential activation of the melanopsin-mediated versus outer photoreceptor-mediated pupil light reflex.

OBJECTIVE: To weight the rod-, cone-, and melanopsin-mediated activation of the retinal ganglion cells, which drive the pupil light reflex by varying the light stimulus wavelength, intensity, and duration. DESIGN: Experimental study. PARTICIPANTS: Forty-three subjects with normal eyes and 3 patients with neuroretinal visual loss. METHODS: A novel stimulus paradigm was developed using either a long wavelength (red) or short wavelength (blue) light given as a continuous Ganzfeld stimulus with stepwise increases over a 2 log-unit range. The pupillary movement before, during, and after the light stimulus was recorded in real time with an infrared illuminated video camera. MAIN OUTCOME MEASURES: The percent pupil contraction of the transient and sustained pupil response to a low- (1 cd/m(2)), medium- (10 cd/m(2)), and high-intensity (100 cd/m(2)) red- and blue-light stimulus was calculated for 1 eye of each subject. From the 43 normal eyes, median and 25th, 75th, 5th, and 95th percentile values were obtained for each stimulus condition. RESULTS: In normal eyes at lower intensities, blue light evoked much greater pupil responses compared with red light when matched for photopic luminance. The transient pupil contraction was generally greater than the sustained contraction, and this disparity was greatest at the lowest light intensity and least apparent with bright (100 cd/m(2)) blue light. A patient with primarily rod dysfunction (nonrecordable scotopic electroretinogram) showed significantly reduced pupil responses to blue light at lower intensities. A patient with achromatopsia and an almost normal visual field showed selective reduction of the pupil response to red-light stimulation. A patient with ganglion cell dysfunction owing to anterior ischemic optic neuropathy demonstrated global loss of pupil responses to red and blue light in the affected eye. CONCLUSIONS: Pupil responses that differ as a function of light intensity and wavelength support the hypothesis that selected stimulus conditions can produce pupil responses that reflect phototransduction primarily mediated by rods, cones, or melanopsin. Use of chromatic pupil responses may be a novel way to diagnose and monitor diseases affecting either the outer or inner retina.

Indoor air quality in a public building following smoking bans

Introduction: Exposure to environmental tobacco smoke (ETS) is a major environmental risk factor. Indoor contaminants come from a variety of sources, which can include inadequate ventilation, volatile organic compounds (VOCs), biological agents, combustion products, and ETS. Because ETS is one of the most frequent causes of IAQ complaints as well as the high mortality of passive smoking, in June 2004 the University of Geneva made the decision to ban smoking inside the so called "Uni-Mail" building, the biggest Swiss University human science building of recent construction, and the ordinance was applied beginning in October 2004. This report presents the finding related to the IAQ of the "Uni-Mail" building before and after smoking bans using nicotine, suspended dust, condensate and PAHs level in air as tracers to perform an assessment of passive tobacco exposure for non-smokers inside the building. Methods: Respirable particles (RSP) A real time aerosol monitor (model DataRAM)was place at sampling post 1, level ground floor. Condensate It consists in extracting any organic matter taken on the glass fibre filters by MeOH, and then measuring the total absorbent of the MeOH extract to the UV wavelength of 447 nm. Nicotine Nicotine was taken by means of cartridges containing of XAD-4 to the fixed flow of 0.5 L/min. The analytical method used for the determination of nicotine is based on gas chromatography with Nitrogen selective detector GC-NPD. Results: Figure 1 shows the box plot density display of 3 parameters before and after smoking bans for all 7 sampling posts: dust, condensate and nicotine in air in μg/m3. Conclusion: Before the smoking ban, the level of the concentrations of respirable particles (RSP) is raised more, average of the day 320 μg/m3, with peaks of more than 1000 μg/m3, compared with the values of the surrounding air between 22 and 30 μg/m3. The nicotine level is definitely more important (average 5.53 μg/m3, field 1.5 to 17.9 μg/m3). Once the smoking bans inside the building were applied, one notes a clear improvement in terms of concentrations of pollutants. For dust, the concentration fell by 3 times (average: 130 μg/m3, range: 40 to 160 μg/m3) and that of nicotine by 10 times (average: 0.53 μg/m3, range: 0 to 1.69 μg/m3) compared to that found before smoking bans. The outdoor air RSP concentration was 22 μg/m3 or 10 times lower. Nicotine seems to be the best tracer for ETS free of interference, independent of location or season.