Putative Origins of Cell-Free DNA in Humans: A Review of Active and Passive Nucleic Acid Release Mechanisms

Through various pathways of cell death, degradation, and regulated extrusion, partial or complete genomes of various origins (e.g., host cells, fetal cells, and infiltrating viruses and microbes) are continuously shed into human body fluids in the form of segmented cell-free DNA (cfDNA) molecules. While the genetic complexity of total cfDNA is vast, the development of progressively efficient extraction, high-throughput sequencing, characterization via bioinformatics procedures, and detection have resulted in increasingly accurate partitioning and profiling of cfDNA subtypes. Not surprisingly, cfDNA analysis is emerging as a powerful clinical tool in many branches of medicine. In addition, the low invasiveness of longitudinal cfDNA sampling provides unprecedented access to study temporal genomic changes in a variety of contexts. However, the genetic diversity of cfDNA is also a great source of ambiguity and poses significant experimental and analytical challenges. For example, the cfDNA population in the bloodstream is heterogeneous and also fluctuates dynamically, differs between individuals, and exhibits numerous overlapping features despite often originating from different sources and processes. Therefore, a deeper understanding of the determining variables that impact the properties of cfDNA is crucial, however, thus far, is largely lacking. In this work we review recent and historical research on active vs. passive release mechanisms and estimate the significance and extent of their contribution to the composition of cfDNA.

Neuropod Cells: The Emerging Biology of Gut-Brain Sensory Transduction

Guided by sight, scent, texture, and taste, animals ingest food. Once ingested, it is up to the gut to make sense of the food's nutritional value. Classic sensory systems rely on neuroepithelial circuits to convert stimuli into signals that guide behavior. However, sensation of the gut milieu was thought to be mediated only by the passive release of hormones until the discovery of synapses in enteroendocrine cells. These are gut sensory epithelial cells, and those that form synapses are referred to as neuropod cells. Neuropod cells provide the foundation for the gut to transduce sensory signals from the intestinal milieu to the brain through fast neurotransmission onto neurons, including those of the vagus nerve. These findings have sparked a new field of exploration in sensory neurobiology—that of gut-brain sensory transduction.

Comparison of four inflation techniques on endotracheal tube cuff pressure using a feline airway simulator

Objectives The aim of this study was to compare four inflation techniques on endotracheal tube cuff (ETC) pressure using a feline airway simulator. Methods Ten participants used four different endotracheal cuff inflation techniques to inflate the cuff of a low-pressure, high-volume endotracheal tube within a feline airway simulator. The simulator replicated an average-sized feline trachea, intubated with a 4.5 mm endotracheal tube, connected to a circle breathing system and pressure-controlled ventilation with oxygen and medical air. Participants inflated the ETC: by pilot balloon palpation (P); by instilling the minimum occlusive volume (MOV) required for loss of airway leaks during mechanical ventilation; until a passive release of pressure with use of a loss-of-resistance syringe (LOR); and with use of a syringe with a digital pressure reader (D) specifically designed for endotracheal cuff inflation. Intracuff pressure was measured by a manometer obscured to participants. The ideal pressure was considered to be between 20 and 30 cmH2O. Data were analysed by Shapiro–Wilk, Kruskal–Wallis and χ2 tests, as appropriate. Results Participants were eight veterinarians and two veterinary nurses with additional training in anaesthesia. Measured median intracuff pressures for P, MOV, LOR and D, respectively, were 25 cmH2O (range 4–74 cmH2O), 41 cmH2O (range 4–70 cmH2O), 31 cmH2O (range 18–64 cmH2O) and 22 cmH2O (range 20–30 cmH2O). D performed significantly better ( P <0.001) than all other techniques, with no difference between the other techniques. Conclusions and relevance Use of D for cuff inflation achieved optimal cuff pressures. There may be high operator-dependent variability in the cuff pressures achieved with the use of P, MOV or LOR inflation techniques. As such, a cuff manometer is recommended when using any of these techniques.

A Low-Cost, Passive Release Device for the Surveillance and Control of Mosquitoes

Mosquitoes continue to be a major threat to global health, and the ability to reliably monitor, catch, and kill mosquitoes via passive traps is of great importance. Global, low-cost, and easy-to-use outdoor devices are needed to augment existing efforts in mosquito control that combat the spread of disease, such as Zika. Thus, we have developed a modular, portable, non-powered (passive), self-contained, and field-deployable device suitable for releasing volatiles with a wide range of applications such as attracting, repelling, and killing mosquitoes. This unique device relies on a novel nested wick and two-reservoir design that achieves a constant release of volatiles over several hundred hours. Devices loaded with one of either two compounds, geraniol or 1-methylpiperazine (MP), were tested in a controlled environment (32 °C and 70% relative humidity), and both compounds achieved a constant release from our devices at a rate of 2.4 mg/h and 47 mg/h, respectively. The liquid payload can be volatile attractants or repellants as well as mosquitocide-containing feeding solutions for capture and surveillance. This low-cost device can be utilized for both civilian and military mosquito control purposes, but it will be particularly important for protecting those in economically repressed environments, such as sub-Saharan Africa and Central and South America.

Hybrid Two-Scale Fabrication of Sub-Millimetric Capillary Grippers

Capillary gripping is a pick-and-place technique that is particularly well-suited for handling sub-millimetric components. Nevertheless, integrating a fluid supply and release mechanism becomes increasingly difficult to manufacture for these scales. In the present contribution, two hybrid manufacturing procedures are introduced in which the creation of the smallest features is decoupled from the macro-scale components. In the first procedure, small scale features are printed directly (by two-photon polymerisation) on top of a 3D-printed device (through stereolithography). In the second approach, directional ultraviolet (UV)-illumination and an adapted design allowed for successful (polydimethylsiloxane, PDMS) moulding of the microscopic gripper head on top of a metal substrate. Importantly, a fully functional microchannel is present in both cases through which liquid to grip the components can be supplied and retracted. This capability of removing the liquid combined with an asymmetric pillar design allows for a passive release mechanism with a placement precision on the order of 3% of the component size.

A gut-brain neural circuit for nutrient sensory transduction

The brain is thought to sense gut stimuli only via the passive release of hormones. This is because no connection has been described between the vagus and the putative gut epithelial sensor cell—the enteroendocrine cell. However, these electrically excitable cells contain several features of epithelial transducers. Using a mouse model, we found that enteroendocrine cells synapse with vagal neurons to transduce gut luminal signals in milliseconds by using glutamate as a neurotransmitter. These synaptically connected enteroendocrine cells are referred to henceforth as neuropod cells. The neuroepithelial circuit they form connects the intestinal lumen to the brainstem in one synapse, opening a physical conduit for the brain to sense gut stimuli with the temporal precision and topographical resolution of a synapse.

Visfatin Is Actively Secreted In Vitro From U-937 Macrophages, but Only Passively Released From 3T3-L1 Adipocytes and HepG2 Hepatocytes

Visfatin is a multi-functional molecule that can act intracellularly and extracellularly as an adipokine, cytokine and enzyme. One of the main questions concerning visfatin is the mechanism of its secretion; whether, how and from which cells visfatin is released. The objective of this in vitro study was to observe the active secretion of visfatin from 3T3-L1 preadipocytes and adipocytes, HepG2 hepatocytes, U-937, THP-1 and HL-60 monocytes and macrophages. The amount of visfatin in media and cell lysate was always related to the intracellular enzyme, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), to exclude the passive release of visfatin. Visfatin was not found in media of 3T3-L1 preadipocytes. In media of 3T3-L1 adipocytes and HepG2 hepatocytes, the ratio of visfatin to the amount of GAPDH was identical to cell lysates. Hence, it is likely that these cells do not actively secrete visfatin in a significant manner. However, we found that significant producers of visfatin are differentiated macrophages and that the amount of secreted visfatin depends on used cell line and it is affected by the mode of differentiation. Results show that 3T3-L1 adipocytes and HepG2 hepatocytes released visfatin only passively during the cell death. U-937 macrophages secrete visfatin in the greatest level from all of the tested cell lines.

Porous PDMS structures for the storage and release of aqueous solutions into fluidic environments

This work introduces a highly porous PDMS sponge for the storage and passive release of aqueous solutions, acting as a building block for self-sufficient microfluidic systems.