Physiological and pathological roles for R-loop structures continue to be discovered, and studies suggest that R-loops could contribute to human disease. R-loops are nucleic acid structures characterized by a DNA:RNA hybrid and displaced single-stranded DNA that occur in connection with transcription. R-loops form naturally and have been shown to be important for a number of physiological processes such as mitochondrial replication initiation, class switch recombination, DNA repair, modulating DNA topology, and regulation of gene expression. However, subsets of R-loops or persistent R-loops lead to DNA breaks, chromosome rearrangement, and genome instability. In addition, R-loops have been linked to human diseases, specifically neurological disorders and cancer. Of the large amount of research produced recently on R-loops, this review covers evidence for R-loop involvement in normal cellular physiology and pathophysiology, as well as describing factors that contribute to R-loop regulation.

Skeletal muscle-specific liver kinase B1 (LKB1) knockout mice (skmLKB1-KO) exhibit elevated mitogen-activated protein kinase (MAPK) signaling after treadmill running. MAPK activation is also associated with inflammation-related signaling in skeletal muscle. Since exercise can induce muscle damage, and inflammation is a response triggered by damaged tissue, we therefore hypothesized that LKB1 plays an important role in dampening the inflammatory response to muscle contraction, and that this may be due in part to increased susceptibility to muscle damage with contractions in LKB1-deficient muscle. Here we studied the inflammatory response and muscle damage with in situ muscle contraction or downhill running. After in situ muscle contractions, the phosphorylation of both NF-κB and STAT3 was increased more in skmLKB1-KO vs. wild-type (WT) muscles. Analysis of gene expression via microarray and RT-PCR shows that expression of many inflammation-related genes increased after contraction only in skmLKB1-KO muscles. This was associated with mild skeletal muscle fiber membrane damage in skmLKB1-KO muscles. Gene markers of oxidative stress were also elevated in skmLKB1-KO muscles after contraction. Using the downhill running model, we observed significantly more muscle damage after running in skmLKB1-KO mice, and this was associated with greater phosphorylation of both Jnk and STAT3 and increased expression of SOCS3 and Fos. In conclusion, we have shown that the lack of LKB1 in skeletal muscle leads to an increased inflammatory state in skeletal muscle that is exacerbated by muscle contraction. Increased susceptibility of the muscle to damage may underlie part of this response.

IntroductionThe COVID-19 pandemic has revealed an immense, unmet and international need for available ventilators. Both clinical and engineering groups around the globe have responded through the development of “homemade” or do-it-yourself (DIY) ventilators. Several designs have been prototyped, tested, and shared over the internet. However, many open source DIY ventilators require extensive familiarity with microcontroller programming and electronics assembly, which many healthcare providers may lack. In light of this, we designed and bench tested a low-cost, pressure-controlled mechanical ventilator that is “plug and play” by design, where no end-user microcontroller programming is required. This Fast-AssembLy COVID-Nineteen (FALCON) emergency prototype ventilator can be rapidly assembled and could be readily modified and improved upon to potentially provide a ventilatory option when no other is present, especially in low- and middle-income countries.HypothesisWe anticipated that a minimal component prototype ventilator could be easily assembled that could reproduce pressure/flow waveforms and tidal volumes similar to a hospital grade ventilator (Engström CarestationTM).Materials and MethodsWe benched-tested our prototype ventilator using an artificial test lung under 36 test conditions with varying respiratory rates, peak inspiratory pressures (PIP), positive end expiratory pressures (PEEP), and artificial lung compliances. Pressure and flow waveforms were recorded, and tidal volumes calculated with prototype ventilator performance compared to a hospital-grade ventilator (Engström CarestationTM) under identical test conditions.ResultsPressure and flow waveforms produced by the prototype ventilator were highly similar to the CarestationTM. The ventilator generated consistent PIP/PEEP, with tidal volume ranges similar to the CarestationTM. The FALCON prototype was tested continuously for a 5-day period without failure or significant changes in delivered PIP/PEEP.ConclusionThe FALCON prototype ventilator is an inexpensive and easily-assembled “plug and play” emergency ventilator design. The FALCON ventilator is currently a non-certified prototype that, following further appropriate validation and testing, might eventually be used as a life-saving emergency device in extraordinary circumstances when more sophisticated forms of ventilation are unavailable.

Anaplastic thyroid cancer (ATC) is one of the most lethal malignancies with a median survival time of about 4 months. Currently, there is no effective treatment, and the development of new therapies is an important and urgent issue for ATC patients. YM155 is a small molecule that was identified as the top candidate in a high-throughput screen of small molecule inhibitors performed against a panel of ATC cell lines by the National Cancer Institute. However, there were no follow-up studies investigating YM155 in ATC. Here, we determined the effects of YM155 on ATC and human primary benign thyroid cell (PBTC) survival with alamarBlue assay. Our data show that YM155 inhibited proliferation of ATC cell lines while sparing normal thyroid cells, suggesting a high therapeutic window. YM155-induced DNA damage was detected by measuring phosphorylation of -H2AX as a marker for DNA double-strand breaks. The formamidopyrimidine-DNA glycosylase (FPG)-modified alkaline comet assay in conjunction with reactive oxygen species (ROS) assay and glutathione (GSH)/glutathione (GSSG) assay suggests that YM155-mediated oxidative stress contributes to DNA damage. In addition, we provide evidence that YM155 causes cell cycle arrest in S phase and in the G2/M transition and causes apoptosis, as seen with flow cytometry. In this study, we show for the first time the multiple effects of YM155 in ATC cells, furthering a potential therapeutic approach for ATC.

Supplementary Figure from YM155 Induces DNA Damage and Cell Death in Anaplastic Thyroid Cancer Cells by Inhibiting DNA Topoisomerase IIα at the ATP-Binding Site