Automated Biochemical, Morphological, and Organizational Assessment of Precancerous Changes from Endogenous Two-Photon Fluorescence Images

Multi-photon fluorescence microscopy techniques allow for non-invasive interrogation of live samples in their native environment. These methods are particularly appealing for identifying pre-cancers because they are sensitive to the early changes that occur on the microscopic scale and can provide additional information not available using conventional screening techniques.In this study, we developed novel automated approaches, which can be employed for the real-time analysis of two-photon fluorescence images, to non-invasively discriminate between normal and pre-cancerous/HPV-immortalized engineered tissues by concurrently assessing metabolic activity, morphology, organization, and keratin localization. Specifically, we found that the metabolic activity was significantly enhanced and more uniform throughout the depths of the HPV-immortalized epithelia, based on our extraction of the NADH and FAD fluorescence contributions. Furthermore, we were able to separate the keratin contribution from metabolic enzymes to improve the redox estimates and to use the keratin localization as a means to discriminate between tissue types. To assess morphology and organization, Fourier-based, power spectral density (PSD) approaches were employed. The nuclear size distribution throughout the epithelial depths was quantified by evaluating the variance of the corresponding spatial frequencies, which was found to be greater in the normal tissue compared to the HPV-immortalized tissues. The PSD was also used to calculate the Hurst parameter to identify the level of organization in the tissues, assuming a fractal model for the fluorescence intensity fluctuations within a field. We found the range of organization was greater in the normal tissue and closely related to the level of differentiation.A wealth of complementary morphological, biochemical and organizational tissue parameters can be extracted from high resolution images that are acquired based entirely on endogenous sources of contrast. They are promising diagnostic parameters for the non-invasive identification of early cancerous changes and could improve significantly diagnosis and treatment for numerous patients

Quercetin elevates p27Kip1 and arrests both primary and HPV16 E6/E7 transformed human keratinocytes in G1

Our previous work with primary bovine fibroblasts demonstrated that quercetin, a potent mutagen found in high levels in bracken fern (Pteridium aquilinum), arrested cells in G1 and G2/M, in correlation with p53 activation. The expression of bovine papillomavirus type 4 (BPV-4) E7 overcame this arrest and lead to the development of tumorigenic cells lines (Beniston et al., 2001). Given the possible link between papillomavirus infection, bracken fern in the diet and cancer of the upper gastrointestinal (GI) tract in humans, we investigated whether a similar situation would occur in human cells transformed by human papillomavirus type 16 (HPV-16) oncoproteins. Quercetin arrested primary human foreskin keratinocytes in G1. Arrest was linked to an elevation of the cyclin-dependent kinase inhibitor (cdki) p27Kip1. Expression of the HPV16 E6 and E7 oncoproteins in transformed cells failed to abrogate cell cycle arrest. G1 arrest in the transformed cells was also linked to an increase of p27Kip1 with a concomitant reduction of cyclin E-associated kinase activity. This elevation of p27Kip1 was due not only to increased protein half-life, but also to increased mRNA transcription

Modulating prospective memory and attentional control with high-definition transcranial current stimulation: Study protocol of a randomized, double-blind, and sham-controlled trial in healthy older adults.

The ability to remember future intentions (i.e., prospective memory) is influenced by attentional control. At the neuronal level, frontal and parietal brain regions have been related to attentional control and prospective memory. It is debated, however, whether more or less activity in these regions is beneficial for older adults' performance. We will test that by systematically enhancing or inhibiting activity in these regions with anodal or cathodal high-definition transcranial direct current stimulation in older adults. We will include n = 105 healthy older volunteers (60-75 years of age) in a randomized, double-blind, sham-controlled, and parallel-group design. The participants will receive either cathodal, anodal, or sham high-definition transcranial direct current stimulation of the left or right inferior frontal gyrus, or the right superior parietal gyrus (1mA for 20 min). During and after stimulation, the participants will complete tasks of attentional control and prospective memory. The results of this study will clarify how frontal and parietal brain regions contribute to attentional control and prospective memory in older healthy adults. In addition, we will elucidate the relationship between attentional control and prospective memory in that age group. The study has been registered with ClinicalTrials.gov on the 12th of May 2021 (trial identifier: NCT04882527)

Genomic monitoring to understand the emergence and spread of Usutu virus in the Netherlands, 2016-2018

Usutu virus (USUV) is a mosquito-borne flavivirus circulating in Western Europe that causes die-offs of mainly common blackbirds (Turdus merula). In the Netherlands, USUV was first detected in 2016, when it was identified as the likely cause of an outbreak in birds. In this study, dead blackbirds were collected, screened for the presence of USUV and submitted to Nanopore-based sequencing. Genomic sequence

Critical Exponents of the Classical 3D Heisenberg Model: A Single-Cluster Monte Carlo Study

We have simulated the three-dimensional Heisenberg model on simple cubic lattices, using the single-cluster Monte Carlo update algorithm. The expected pronounced reduction of critical slowing down at the phase transition is verified. This allows simulations on significantly larger lattices than in previous studies and consequently a better control over systematic errors. In one set of simulations we employ the usual finite-size scaling methods to compute the critical exponents $\nu,\alpha,\beta,\gamma, \eta$ from a few measurements in the vicinity of the critical point, making extensive use of histogram reweighting and optimization techniques. In another set of simulations we report measurements of improved estimators for the spatial correlation length and the susceptibility in the high-temperature phase, obtained on lattices with up to $100^3$ spins. This enables us to compute independent estimates of $\nu$ and $\gamma$ from power-law fits of their critical divergencies.Comment: 33 pages, 12 figures (not included, available on request). Preprint FUB-HEP 19/92, HLRZ 77/92, September 199