Real-Time Intrinsic Fluorescence Visualization and Sizing of Proteins and Protein Complexes in Microfluidic Devices.

Optical detection has become a convenient and scalable approach to read out information from microfluidic systems. For the study of many key biomolecules, however, including peptides and proteins, which have low fluorescence emission efficiencies at visible wavelengths, this approach typically requires labeling of the species of interest with extrinsic fluorophores to enhance the optical signal obtained - a process which can be time-consuming, requires purification steps, and has the propensity to perturb the behavior of the systems under study due to interactions between the labels and the analyte molecules. As such, the exploitation of the intrinsic fluorescence of protein molecules in the UV range of the electromagnetic spectrum is an attractive path to allow the study of unlabeled proteins. However, direct visualization using 280 nm excitation in microfluidic devices has to date commonly required the use of coherent sources with frequency multipliers and devices fabricated out of materials that are incompatible with soft lithography techniques. Here, we have developed a simple, robust, and cost-effective 280 nm LED platform that allows real-time visualization of intrinsic fluorescence from both unlabeled proteins and protein complexes in polydimethylsiloxane microfluidic channels fabricated through soft lithography. Using this platform, we demonstrate intrinsic fluorescence visualization of proteins at nanomolar concentrations on chip and combine visualization with micron-scale diffusional sizing to measure the hydrodynamic radii of individual proteins and protein complexes under their native conditions in solution in a label-free manner

Second-order L2L^2-regularity in nonlinear elliptic problems

A second-order regularity theory is developed for solutions to a class of quasilinear elliptic equations in divergence form, including the $p$-Laplace equation, with merely square-integrable right-hand side. Our results amount to the existence and square integrability of the weak derivatives of the nonlinear expression of the gradient under the divergence operator. This provides a nonlinear counterpart of the classical $L^2$-coercivity theory for linear problems, which is missing in the existing literature. Both local and global estimates are established. The latter apply to solutions to either Dirichlet or Neumann boundary value problems. Minimal regularity on the boundary of the domain is required. If the domain is convex, no regularity of its boundary is needed at all

Risk of Childhood Cancers Associated with Residence in Agriculturally Intense Areas in the United States

Background: The potential for widespread exposure to agricultural pesticides through drift during application raises concerns about possible health effects to exposed children living in areas of high agricultural activity. Objectives: We evaluated whether residence in a county with greater agricultural activity was associated with risk of developing cancer in children \u3c 15 years of age. Methods: Incidence data for U.S. children 0–14 years of age diagnosed with cancer between 1995 and 2001 were provided by member registries of the North American Association of Central Cancer Registries. We determined percent cropland for each county using agricultural census data, and used the overall study distribution to classify agriculturally intense counties. We estimated odds ratios and 95% confidence intervals for all ages and 5-year age groups for total cancers and selected cancer sites using logistic regression. Results: Our study results showed statistically significant increased risk estimates for many types of childhood cancers associated with residence at diagnosis in counties having a moderate to high level of agricultural activity, with a remarkably consistent dose–response effect seen for counties having ≥ 60% of the total county acreage devoted to farming. Risk for different cancers varied by type of crop. Conclusions: Although interpretation is limited by the ecologic design, in this study we were able to evaluate rarer childhood cancers across a diverse agricultural topography. The findings of this exploratory study support a continued interest in the possible impact of long-term, low-level pesticide exposure in communities located in agriculturally intense areas

Evidence for weathering and volcanism during the PETM from Arctic Ocean and Peri-Tethys osmium isotope records

Sudden global warming during the Paleocene–Eocene Thermal Maximum (PETM, 55.9 Ma) occurred because of the rapid release of several thousand gigatonnes of isotopically light carbon into the oceans and atmosphere; however, the cause of this release is not well understood. Some studies have linked carbon injection to volcanic activity associated with the North Atlantic Igneous Province (NAIP), while others have emphasised carbon cycle feedbacks associated with orbital forcing. This study presents the osmium isotope compositions of mudrocks that were deposited during the PETM at four locations (one from the Arctic Ocean, and three from the Peri-Tethys). The Os-isotope records all exhibit a shift of similar magnitude towards relatively radiogenic values across the PETM. This observation confirms that there was a transient, global increase in the flux of radiogenic Os from the weathering of continental rocks in response to elevated temperatures at that time. The tectonic effects of NAIP volcanic emplacement near the onset of the PETM is recorded by anomalously radiogenic Os-isotope compositions of PETM-age Arctic Ocean samples, which indicate an interval of hydrographic restriction that can be linked tectonic uplift due to hotspot volcanism in the North Atlantic seaway. The Peri-Tethys data also document a transient, higher flux of unradiogenic osmium into the ocean near the beginning of the PETM, most likely from the weathering of young mafic rocks associated with the NAIP. These observations support the hypothesis that volcanism played a major role in triggering the cascade of environmental changes during the PETM, and highlight the influence of paleogeography on the Os isotope characteristics of marine water masses

Structure-Guided Optimization of Replication Protein A (RPA)–DNA Interaction Inhibitors

Replication protein A (RPA) is the major human single stranded DNA (ssDNA)-binding protein, playing essential roles in DNA replication, repair, recombination, and DNA-damage response (DDR). Inhibition of RPA–DNA interactions represents a therapeutic strategy for cancer drug discovery and has great potential to provide single agent anticancer activity and to synergize with both common DNA damaging chemotherapeutics and newer targeted anticancer agents. In this letter, a new series of analogues based on our previously reported TDRL-551 (4) compound were designed to improve potency and physicochemical properties. Molecular docking studies guided molecular insights, and further SAR exploration led to the identification of a series of novel compounds with low micromolar RPA inhibitory activity, increased solubility, and excellent cellular up-take. Among a series of analogues, compounds 43, 44, 45, and 46 hold promise for further development of novel anticancer agents

Design and Structure-Guided Development of Novel Inhibitors of the Xeroderma Pigmentosum Group A (XPA) Protein–DNA Interaction

XPA is a unique and essential protein required for the nucleotide excision DNA repair pathway and represents a therapeutic target in oncology. Herein, we are the first to develop novel inhibitors of the XPA–DNA interaction through structure-guided drug design efforts. Ester derivatives of the compounds 1 (X80), 22, and 24 displayed excellent inhibitory activity (IC50 of 0.82 ± 0.18 μM and 1.3 ± 0.22 μM, respectively) but poor solubility. We have synthesized novel amide derivatives that retain potency and have much improved solubility. Furthermore, compound 1 analogs exhibited good specificity for XPA over RPA (replication protein A), another DNA-binding protein that participates in the nucleotide excision repair (NER) pathway. Importantly, there were no significant interactions observed by the X80 class of compounds directly with DNA. Molecular docking studies revealed a mechanistic model for the interaction, and these studies could serve as the basis for continued analysis of structure–activity relationships and drug development efforts of this novel target

Multidisciplinary approach to reconstructing local pastoral activities: an example from the Pyrenean Mountains (Pays Basque)

International audienceIn this study archaeology, history and palaeoecology (modern and fossil data sets of pollen and nonpollen palynomorphs) were used to reconstruct small-scale pastoral activities in the Pyrenees Mountains during the last two millennia. Modern pollen assemblages from the major vegetation units (both natural andanthropogenic) are studied on one restricted watershed area. A correlative model (RDA) of 61 modern pollen spectra and 35 external variables distinguishes two groups of taxa, providing information on the nature and spatial extent of human impact on the landscape. The first pool indicates local pastoral activities, and the second one implies regional input from outside the studied watershed, and is not characteristic of a specific land use. These pools are described as 'Local Pastoral Pollen Indicators' (LPPI) for this particular mountain region on crystalline bedrock and 'Regional Human Activities Pollen Indicators' (RHAPI). The modern data set is used to aid interpretation of the local pollen sequence of Sourzay that covers the last 2000 calendar years BP, using RDA reconstructions, and best modern analogues as a means of comparing modern and fossil spectra. The study also demonstrates agreement between the independent interpretations of two fossil proxies, LPPI and coprophilous fungi

A public early intervention approach to first-episode psychosis: Treated incidence over 7 years in the Emilia-Romagna region

AimTo estimate the treated incidence of individuals with first-episode psychosis (FEP) who contacted the Emilia-Romagna public mental healthcare system (Italy); to examine the variability of incidence and user characteristics across centres and years. MethodsWe computed the raw treated incidence in 2013-2019, based on FEP users aged 18-35, seen within or outside the regional program for FEP. We modelled FEP incidence across 10 catchment areas and 7 years using Bayesian Poisson and Negative Binomial Generalized Linear Models of varying complexity. We explored associations between user characteristics, study centre and year comparing variables and socioclinical clusters of subjects. ResultsThousand three hundred and eighteen individuals were treated for FEP (raw incidence: 25.3 / 100.000 inhabitant year, IQR: 15.3). A Negative Binomial location-scale model with area, population density and year as predictors found that incidence and its variability changed across centres (Bologna: 36.55; 95% CrI: 30.39-43.86; Imola: 3.07; 95% CrI: 1.61-4.99) but did not follow linear temporal trends or density. Centers were associated with different user age, gender, migrant status, occupation, living conditions and cluster distribution. Year was associated negatively with HoNOS score (R = -0.09, p < .001), duration of untreated psychosis (R = -0.12, p < .001) and referral type. ConclusionsThe Emilia-Romagna region presents a relatively high but variable incidence of FEP across areas, but not in time. More granular information on social, ethnic and cultural factors may increase the level of explanation and prediction of FEP incidence and characteristics, shedding light on social and healthcare factors influencing FEP