Lipids and Proteins Differentiation in Membrane Fouling Using Heavy Metal Staining and Electron Microscopy at Cryogenic Temperatures.

The detailed characterization of fouling in membranes is essential to understand any observed improvement or reduction on filtration performance. Electron microscopy allows detailed structural characterization, and its combination with labeling techniques, using electron-dense probes, typically allows for the differentiation of biomolecules. Developing specific protocols that allow for differentiation of biomolecules in membrane fouling by electron microscopy is a major challenge due to both as follows: the necessity to preserve the native state of fouled membranes upon real filtration conditions as well as the inability of the electron-dense probes to penetrate the membranes once they have been fouled. In this study, we present the development of a heavy metal staining technique for identification and differentiation of biomolecules in membrane fouling, which is compatible with cryofixation methods. A general contrast enhancement of biomolecules and fouling is achieved. Our observations indicate a strong interaction between biomolecules: A tendency of proteins, both in solution as well as in the fouling, to surround the lipids is observed. Using transmission electron microscopy and scanning electron microscopy at cryogenic conditions, cryo-SEM, in combination with energy-dispersive X-ray spectroscopy, the spatial distribution of proteins and lipids within fouling is shown and the role of proteins in fouling discussed.

Localizing and lateralizing value of auditory phenomena in seizures.

BACKGROUND: Auditory seizures (AS) are a rare type of focal seizures. AS are classically thought to involve a seizure onset zone (SOZ) in the temporal lobe, but there remain uncertainties about their localizing and lateralizing value. We conducted a narrative literature review with the aim of providing an up-to-date description of the lateralizing and localizing value of AS. METHODS: The databases PubMed, Scopus, and Google Scholar were searched for literature on AS in December 2022. All cortical stimulation studies, case reports, and case series were analyzed to assess for auditory phenomena that were suggestive of AS and to evaluate if the lateralization and/or localization of the SOZ could be determined. We classified AS according to their semiology (e.g., simple hallucination versus complex hallucination) and the level of evidence with which the SOZ could be predicted. RESULTS: A total of 174 cases comprising 200 AS were analyzed from 70 articles. Across all studies, the SOZ of AS were more often in the left (62%) than in the right (38%) hemisphere. AS heard bilaterally followed this trend. Unilaterally heard AS were more often due to a SOZ in the contralateral hemisphere (74%), although they could also be ipsilateral (26%). The SOZ for AS was not limited to the auditory cortex, nor to the temporal lobe. The areas more frequently involved in the temporal lobe were the superior temporal gyrus (STG) and mesiotemporal structures. Extratemporal locations included parietal, frontal, insular, and rarely occipital structures. CONCLUSION: Our review highlighted the complexity of AS and their importance in the identification of the SOZ. Due to the limited data and heterogeneous presentation of AS in the literature, the patterns associated with different AS semiologies warrant further research.

Sudden unexpected death in epilepsy (SUDEP): A bibliometric analysis.

OBJECTIVE: The literature on sudden unexpected death in epilepsy (SUDEP) has been evolving at a staggering rate. We conducted a bibliometric analysis of the SUDEP literature with the aim of presenting its structure, performance, and trends. METHODS: The Scopus database was searched in April 2023 for documents explicitly detailing SUDEP in their title, abstract, or keywords. After the removal of duplicate documents, bibliometric analysis was performed using the R package bibliometrix and the program VOSviewer. Performance metrics were computed to describe the literature's annual productivity, most relevant authors and countries, and most important publications. Science mapping was performed to visualize the relationships between research constituents by constructing a country collaboration network, co-authorship network, keyword co-occurrence network, and document co-citation network. RESULTS: A total of 2140 documents were analyzed. These documents were published from 1989 onward, with an average number of citations per document of 25.78. Annual productivity had been on the rise since 2006. Out of 6502 authors, five authors were in both the list of the ten most productive and the list of the ten most cited authors: Devinsky O, Sander JW, Tomson T, Ryvlin P, and Lhatoo SD. The USA and the United Kingdom were the most productive and cited countries. Collaborations between American authors and European authors were particularly rich. Prominent themes in the literature included those related to pathophysiology (e.g., cardiac arrhythmia, apnea, autonomic dysfunction), epilepsy characteristics (e.g., epilepsy type, refractoriness, antiseizure medications), and epidemiology (e.g., incidence, age, sex). Emerging themes included sleep, genetics, epilepsy refractoriness, and non-human studies. SIGNIFICANCE: The body of literature on SUDEP is rich, fast-growing, and benefiting from frequent international collaborations. Some research themes such as sleep, genetics, and animal studies have become more prevalent over recent years.

Guanine Nucleotide-Binding Protein-Like 1 (GNL1) binds RNA G-quadruplex structures in genes associated with Parkinson's disease.

RNAs are highly regulated at the post-transcriptional level in neurodegenerative diseases and just a few mutations can significantly affect the fate of neuronal cells. To date, the impact of G-quadruplex (G4) regulation in neurodegenerative diseases like Parkinson's disease (PD) has not been analysed. In this study, in silico potential G4s located in deregulated genes related to the nervous system were initially identified and were found to be significantly enriched. Several G4 sequences found in the 5' untranslated regions (5'UTR) of mRNAs associated with Parkinson's disease were demonstrated to in fact fold in vitro by biochemical assays. Subcloning of the full-length 5'UTRs of these candidates upstream of a luciferase reporter system led to the demonstration that the G4s of both Parkin RBR E3 Ubiquitin Protein Ligase (PRKN) and Vacuolar Protein Sorting-Associated Protein 35 (VPS35) significantly repressed the translation of both genes in SH-SY5Y cells. Subsequently, a strategy of using label-free RNA affinity purification assays with either of these two G4 sequences as bait isolated the Guanine Nucleotide-Binding Protein-Like 1 (GNL1). The latter was shown to have a higher affinity for the G4 sequences than for their mutated version. This study sheds light on new RNA G-quadruplexes located in genes dysregulated in Parkinson disease and a new G4-binding protein, GNL1.

Investigating Tunneling Nanotubes in Cancer Cells: Guidelines for Structural and Functional Studies through Cell Imaging.

By allowing insured communication between cancer cells themselves and with the neighboring stromal cells, tunneling nanotubes (TNTs) are involved in the multistep process of cancer development from tumorigenesis to the treatment resistance. However, despite their critical role in the biology of cancer, the study of the TNTs has been announced challenging due to not only the absence of a specific biomarker but also the fragile and transitory nature of their structure and the fact that they are hovering freely above the substratum. Here, we proposed to review guidelines to follow for studying the structure and functionality of TNTs in tumoral neuroendocrine cells (PC12) and nontumorigenic human bronchial epithelial cells (HBEC-3, H28). In particular, we reported how crucial is it (i) to consider the culture conditions (culture surface, cell density), (ii) to visualize the formation of TNTs in living cells (mechanisms of formation, 3D representation), and (iii) to identify the cytoskeleton components and the associated elements (categories, origin, tip, and formation/transport) in the TNTs. We also focused on the input of high-resolution cell imaging approaches including Stimulated Emission Depletion (STED) nanoscopy, Transmitted and Scanning Electron Microscopies (TEM and SEM). In addition, we underlined the important role of the organelles in the mechanisms of TNT formation and transfer between the cancer cells. Finally, new biological models for the identification of the TNTs between cancer cells and stromal cells (liquid air interface, ex vivo, in vivo) and the clinical considerations will also be discussed.

A role for RASSF1A in tunneling nanotube formation between cells through GEFH1/Rab11 pathway control.

BACKGROUND: By allowing intercellular communication between cells, tunneling nanotubes (TNTs) could play critical role in cancer progression. If TNT formation is known to require cytoskeleton remodeling, key mechanism controlling their formation remains poorly understood. METHODS: The cells of human bronchial (HBEC-3, A549) or mesothelial (H2452, H28) lines are transfected with different siRNAs (inactive, anti-RASSF1A, anti-GEFH1 and / or anti-Rab11). At 48 h post-transfection, i) the number and length of the nanotubes per cell are quantified, ii) the organelles, previously labeled with specific tracers, exchanged via these structures are monitored in real time between cells cultured in 2D or 3D and in normoxia, hypoxia or in serum deprivation condition. RESULTS: We report that RASSF1A, a key-regulator of cytoskeleton encoded by a tumor-suppressor gene on 3p chromosome, is involved in TNTs formation in bronchial and pleural cells since controlling proper activity of RhoB guanine nucleotide exchange factor, GEF-H1. Indeed, the GEF-H1 inactivation induced by RASSF1A silencing, leads to Rab11 accumulation and subsequent exosome releasing, which in turn contribute to TNTs formation. Finally, we provide evidence involving TNT formation in bronchial carcinogenesis, by reporting that hypoxia or nutriment privation, two almost universal conditions in human cancers, fail to prevent TNTs induced by the oncogenic RASSF1A loss of expression. CONCLUSIONS: This finding suggests for the first time that loss of RASSF1A expression could be a potential biomarker for TNTs formation, such TNTs facilitating intercellular communication favoring multistep progression of bronchial epithelial cells toward overt malignancy.

Population health impact of cancer in Canada, 2001.

Summary measures of population health that incorporate morbidity provide a new perspective for health policy and priority setting. Health-adjusted life years (HALYs) lost to a disease combine the impact of years of life lost to premature mortality and morbidity, measured as year-equivalents lost to reduced functioning. HALYs for 25 cancers were estimated from mortality and incidence in 2001 in Canada; population-attributable fractions were estimated for major risk factors contributing to these cancers. Results from this analysis indicate that Canadians would lose an estimated 905,000 health-adjusted years of life to cancer for 2001, including 771,000 to premature mortality and 134,000 to morbidity from incident cases (years discounted at 3 percent). Most of the estimated premature mortality was due to lung cancer; morbidity was largely due to breast, prostate and colorectal cancers. An estimated one quarter of HALYs lost to cancer were attributable to smoking and almost one quarter were attributable to alcohol consumption, lack of fruit and vegetables, obesity and physical inactivity combined. These results are a significant advance in measuring the population health impact of cancer in Canada because they incorporate morbidity as well as mortality.

New application of AAO template: a mold for nanoring and nanocone arrays.

Nanoring and nanocone arrays were prepared by conical openings of nanopores in as-prepared AAO film as a mold.

High fidelity, high yield production of microfluidic devices by hot embossing lithography: rheology and stiction.

We discuss thermoforming of thermoplastic polymers for the hot-embossing lithographic (HEL) fabrication of microfluidic chips near equilibrium conditions that minimize elastic recoil for optimal motif replication. While HEL is often simplistically described as the transfer of micro- and nano-motifs into heat-softened thermoplastic materials, we describe our rational approach to selecting appropriate processing parameters.

Chemical force microscopy for hot-embossing lithography release layer characterization.

We employed variable temperature chemical force microscopy (VT-CFM) using tips silanized with four different hydro- and hydrofluoroalkyl self-assembling monolayers (SAMs) interacting with a thin-film of poly(cyclic olefin), (PCO) to model the hot-embossing stamp-polymer interaction over a temperature range spanning the glass transition of the PCO.

Transition of MEMS technology to nanofabrication.

The transition of MEMS technology to nano fabrication is a solution to the growing demand for smaller and high-density feature sizes in the nanometer scale. Nanoimprint lithography (NIL) techniques for fabricating micro- and nano-features are discussed including hot embossing lithography (HEL), UV Molding (UVM) and micro contact printing (microCP). Recent results in micro and nano-pattern transfer are presented where features ranged from < 100 nm to several centimeters. We also present a comparative study between standard glass microfluidic chips and their HEL counterparts by metrology. Hot-embossed microfluidic chips are shown to be faithful replicates of their parent stamps. NIL is presented as a promising avenue for low-cost, high throughput micro and nano-device fabrication.