Abstract
Background
Although the characterization of cell-free extrachromosomal circular DNA (eccDNA) has gained much research interest, the methylation status of these molecules is yet to be ...elucidated. We set out to compare the methylation densities of plasma eccDNA of maternal and fetal origins, and between small and large molecules. The clearance of fetal eccDNA from maternal circulation was also investigated.
Methods
We developed a sequencing protocol for eccDNA methylation analysis using tagmentation and enzymatic conversion approaches. A restriction enzyme-based approach was applied to verify the tagmentation results. The efficiency of cell-free fetal eccDNA clearance was investigated by fetal eccDNA fraction evaluations at various postpartum time points.
Results
The methylation densities of fetal eccDNA (median: 56.3%; range: 40.5–67.6%) were lower than the maternal eccDNA (median: 66.7%; range: 56.5–75.7%) (P = 0.02, paired t-test). In addition, eccDNA molecules from the smaller peak cluster (180–230 bp) were of lower methylation levels than those from the larger peak cluster (300–450 bp). Both of these findings were confirmed using the restriction enzyme approach. We also observed comparable methylation densities between linear and eccDNA of both maternal and fetal origins. The average half-lives of fetal linear and eccDNA in the maternal blood were 30.2 and 29.7 min, respectively.
Conclusions
We found that fetal eccDNA in plasma was relatively hypomethylated compared to the maternal eccDNA. The methylation densities of eccDNA were positively correlated with their sizes. In addition, fetal eccDNA was found to be rapidly cleared from the maternal blood after delivery, similar to fetal linear DNA.
•There is a need for processed-based biogeochemical models that can simulate monitored parameters for blue carbon project development as burdensome in-situ monitoring requirements currently make many ...blue carbon projects cost-prohibitive.•Here we review the MEM, PEPRMT, DNDC, DayCent and FVS models.•Only the DNDC model can fully account for all necessary parameters.•The DayCent and the MEM/PEPRMT combined models may become useful with further refinement.•Successful application of such models will address one of the biggest barriers to landscape-scale blue carbon project development.
Processed-based biogeochemical mathematical models are powerful tools that are increasingly being used to estimate potential carbon sequestration and greenhouse gas (GHG) impacts of management at a landscape level. These models can simulate some or all of the processes responsible for carbon sequestration and GHG emissions, which can relieve some of the burdensome in-situ monitoring requirements that make many blue carbon projects cost-prohibitive. Here we selectively review five publicly available and widely used biogeochemical models (MEM, PEPRMT, DNDC, DayCent and FVS) including their current applications and limitations towards blue carbon project development. Of the five models, only the DNDC model can be applied to fully account for net sequestration as applicable to blue carbon offset methodologies. With further development, the DayCent and the combined MEM/PEPRMT models may prove to be applicable. Successful application of such models will address one of the biggest barriers to landscape-scale blue carbon project development.
The transcription factor KLF4 functions in post-mitotic epithelial cells to promote differentiation, and functions in a context-dependent fashion as an oncogene. In the skin KLF4 is co-expressed with ...the nuclear receptors RARγ and RXRα, and knockout studies identify formation of the skin permeability barrier as a common function of these three proteins. We utilized a KLF4-transgenic mouse model of skin cancer in combination with cultured epithelial cells to examine functional interactions between KLF4 and retinoic acid receptors. In cultured cells, activation of a conditional, KLF4-estrogen receptor fusion protein by 4-hydroxytamoxifen resulted in rapid upregulation of transcripts for nuclear receptors including RARγ and RXRα. We tested retinoids in epithelial cell transformation assays, including an RAR-selective agonist (all-trans RA), an RXR-selective agonist (9-cis UAB30, rexinoid), and a pan agonist (9-cis RA). Unlike for several other genes, transformation by KLF4 was inhibited by each retinoid, implicating distinct nuclear receptor heterodimers as modulators of KLF4 transforming activity. When RXRα expression was suppressed by RNAi in cultured cells, transformation was promoted and the inhibitory effect of 9-cis UAB30 was attenuated. Similarly as shown for other mouse models of skin cancer, rexinoid prevented skin tumor initiation resulting from induction of KLF4 in basal keratinocytes. Rexinoid permitted KLF4 expression and KLF4-induced cell cycling, but attenuated the KLF4-induced misexpression of cytokeratin 1 in basal cells. Neoplastic lesions including hyperplasia, dysplasia, and squamous cell carcinoma-like lesions were prevented for up to 30 days. Taken together, the results identify retinoid receptors including RXRα as ligand-dependent inhibitors of KLF4-mediated transformation or tumorigenesis.
Gli family members mediate constitutive Hedgehog signaling in the common skin cancer, basal cell carcinoma (BCC). Snail/Snai1 is rapidly induced by Gli1 in vitro, and is coexpressed with Gli1 in ...human hair follicles and skin tumors. In the current study, we generated a dominant-negative allele of Snail, SnaZFD, composed of the zinc-finger domain and flanking sequence. In promoter-reporter assays, SnaZFD blocked the activity of wild-type Snail on the E-cadherin promoter. Snail loss-of-function mediated by SnaZFD or by one of several short hairpin RNAs inhibited transformation of RK3E epithelial cells by Gli1. Conversely, enforced expression of Snail promoted transformation in vitro by Gli1, but not by other genes that were tested, including Notch1, ErbB2, and N-Ras. As observed for Gli1, wild-type Snail repressed E-cadherin in RK3E cells and induced blebbing of the cytoplasmic membrane. Induction of a conditional Gli1 transgene in the basal keratinocytes of mouse skin led to rapid upregulation of Snail transcripts and to cell proliferation in the interfollicular epidermis. Established Gli1-induced skin lesions exhibited molecular similarities to BCC, including loss of E-cadherin. The results identify Snail as a Gli1-inducible effector of transformation in vitro, and an early Gli1-responsive gene in the skin.
Cell-free extrachromosomal circular DNA (eccDNA) as a distinct topological form from linear DNA has recently gained increasing research interest, with possible clinical applications as a class of ...biomarkers. In this study, we aimed to explore the relationship between nucleases and eccDNA characteristics in plasma. By using knockout mouse models with deficiencies in deoxyribonuclease 1 (DNASE1) or deoxyribonuclease 1 like 3 (DNASE1L3), we found that cell-free eccDNA in Dnase1l3-/- mice exhibited larger size distributions than that in wild-type mice. Such size alterations were not found in tissue eccDNA of either Dnase1-/- or Dnase1l3-/- mice, suggesting that DNASE1L3 could digest eccDNA extracellularly but did not seem to affect intracellular eccDNA. Using a mouse pregnancy model, we observed that in Dnase1l3-/- mice pregnant with Dnase1l3+/- fetuses, the eccDNA in the maternal plasma was shorter compared with that of Dnase1l3-/- mice carrying Dnase1l3-/- fetuses, highlighting the systemic effects of circulating fetal DNASE1L3 degrading the maternal eccDNA extracellularly. Furthermore, plasma eccDNA in patients with DNASE1L3 mutations also exhibited longer size distributions than that in healthy controls. Taken together, this study provided a hitherto missing link between nuclease activity and the biological manifestations of eccDNA in plasma, paving the way for future biomarker development of this special form of DNA molecules.
Decisions to continue or suspend therapy with immune checkpoint inhibitors are commonly guided by tumor dynamics seen on serial imaging. However, immunotherapy responses are uniquely challenging to ...interpret because tumors often shrink slowly or can appear transiently enlarged due to inflammation. We hypothesized that monitoring tumor cell death in real time by quantifying changes in circulating tumor DNA (ctDNA) levels could enable early assessment of immunotherapy efficacy.
We compared longitudinal changes in ctDNA levels with changes in radiographic tumor size and with survival outcomes in 28 patients with metastatic non-small cell lung cancer (NSCLC) receiving immune checkpoint inhibitor therapy. CtDNA was quantified by determining the allele fraction of cancer-associated somatic mutations in plasma using a multigene next-generation sequencing assay. We defined a ctDNA response as a >50% decrease in mutant allele fraction from baseline, with a second confirmatory measurement.
Strong agreement was observed between ctDNA response and radiographic response (Cohen's kappa, 0.753). Median time to initial response among patients who achieved responses in both categories was 24.5 days by ctDNA versus 72.5 days by imaging. Time on treatment was significantly longer for ctDNA responders versus nonresponders (median, 205.5 vs. 69 days;
< 0.001). A ctDNA response was associated with superior progression-free survival hazard ratio (HR), 0.29; 95% CI, 0.09-0.89;
= 0.03, and superior overall survival (HR, 0.17; 95% CI, 0.05-0.62;
= 0.007).
A drop in ctDNA level is an early marker of therapeutic efficacy and predicts prolonged survival in patients treated with immune checkpoint inhibitors for NSCLC.
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Provision of safe drinking water in the United States is a great public health achievement. However, new waterborne disease challenges have emerged (e.g., aging infrastructure, chlorine-tolerant and ...biofilm-related pathogens, increased recreational water use). Comprehensive estimates of the health burden for all water exposure routes (ingestion, contact, inhalation) and sources (drinking, recreational, environmental) are needed. We estimated total illnesses, emergency department (ED) visits, hospitalizations, deaths, and direct healthcare costs for 17 waterborne infectious diseases. About 7.15 million waterborne illnesses occur annually (95% credible interval CrI 3.88 million-12.0 million), results in 601,000 ED visits (95% CrI 364,000-866,000), 118,000 hospitalizations (95% CrI 86,800-150,000), and 6,630 deaths (95% CrI 4,520-8,870) and incurring US $3.33 billion (95% CrI 1.37 billion-8.77 billion) in direct healthcare costs. Otitis externa and norovirus infection were the most common illnesses. Most hospitalizations and deaths were caused by biofilm-associated pathogens (nontuberculous mycobacteria, Pseudomonas, Legionella), costing US $2.39 billion annually.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, ODKLJ, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Seismic observations suggest that the uppermost region of Earth’s liquid outer core is buoyant, with slower velocities than the bulk outer core. One possible mechanism for the formation of a stably ...stratified layer is immiscibility in molten iron alloy systems, which has yet to be demonstrated at core pressures. We find immiscibility between liquid Fe-Si and Fe-Si-O persisting to at least 140 GPa through a combination of laser-heated diamond-anvil cell experiments and first-principles molecular dynamics simulations. High-pressure immiscibility in the Fe-Si-O system may explain a stratified layer atop the outer core, complicate differentiation and evolution of the deep Earth, and affect the structure and intensity of Earth’s magnetic field. Our results support silicon and oxygen as coexisting light elements in the core and suggest that SiO₂ does not crystallize out of molten Fe-Si-O at the core-mantle boundary.
Post-COVID-19 condition (also known as long COVID) is an emerging chronic illness potentially affecting millions of people. We aimed to evaluate whether outpatient COVID-19 treatment with metformin, ...ivermectin, or fluvoxamine soon after SARS-CoV-2 infection could reduce the risk of long COVID.
We conducted a decentralised, randomised, quadruple-blind, parallel-group, phase 3 trial (COVID-OUT) at six sites in the USA. We included adults aged 30–85 years with overweight or obesity who had COVID-19 symptoms for fewer than 7 days and a documented SARS-CoV-2 positive PCR or antigen test within 3 days before enrolment. Participants were randomly assigned via 2 × 3 parallel factorial randomisation (1:1:1:1:1:1) to receive metformin plus ivermectin, metformin plus fluvoxamine, metformin plus placebo, ivermectin plus placebo, fluvoxamine plus placebo, or placebo plus placebo. Participants, investigators, care providers, and outcomes assessors were masked to study group assignment. The primary outcome was severe COVID-19 by day 14, and those data have been published previously. Because the trial was delivered remotely nationwide, the a priori primary sample was a modified intention-to-treat sample, meaning that participants who did not receive any dose of study treatment were excluded. Long COVID diagnosis by a medical provider was a prespecified, long-term secondary outcome. This trial is complete and is registered with ClinicalTrials.gov, NCT04510194.
Between Dec 30, 2020, and Jan 28, 2022, 6602 people were assessed for eligibility and 1431 were enrolled and randomly assigned. Of 1323 participants who received a dose of study treatment and were included in the modified intention-to-treat population, 1126 consented for long-term follow-up and completed at least one survey after the assessment for long COVID at day 180 (564 received metformin and 562 received matched placebo; a subset of participants in the metformin vs placebo trial were also randomly assigned to receive ivermectin or fluvoxamine). 1074 (95%) of 1126 participants completed at least 9 months of follow-up. 632 (56·1%) of 1126 participants were female and 494 (43·9%) were male; 44 (7·0%) of 632 women were pregnant. The median age was 45 years (IQR 37–54) and median BMI was 29·8 kg/m2 (IQR 27·0–34·2). Overall, 93 (8·3%) of 1126 participants reported receipt of a long COVID diagnosis by day 300. The cumulative incidence of long COVID by day 300 was 6·3% (95% CI 4·2–8·2) in participants who received metformin and 10·4% (7·8–12·9) in those who received identical metformin placebo (hazard ratio HR 0·59, 95% CI 0·39–0·89; p=0·012). The metformin beneficial effect was consistent across prespecified subgroups. When metformin was started within 3 days of symptom onset, the HR was 0·37 (95% CI 0·15–0·95). There was no effect on cumulative incidence of long COVID with ivermectin (HR 0·99, 95% CI 0·59–1·64) or fluvoxamine (1·36, 0·78–2·34) compared with placebo.
Outpatient treatment with metformin reduced long COVID incidence by about 41%, with an absolute reduction of 4·1%, compared with placebo. Metformin has clinical benefits when used as outpatient treatment for COVID-19 and is globally available, low-cost, and safe.
Parsemus Foundation; Rainwater Charitable Foundation; Fast Grants; UnitedHealth Group Foundation; National Institute of Diabetes, Digestive and Kidney Diseases; National Institutes of Health; and National Center for Advancing Translational Sciences.
Two series of aromatic compounds with perfluoroalkyl (RF) groups of increasing length, 1,3,5,7‐naphthalene(RF)4 and 1,3,5,7,9‐corannulene(RF)5, have been prepared and their electronic properties ...studied by low‐temperature photoelectron spectroscopy (PES) (for gas‐phase electron affinity measurements). These and many related compounds were also studied by DFT calculations. The data demonstrate unambiguously that the electron‐withdrawing ability of RF substituents increases significantly and uniformly from CF3 to C2F5 to n‐C3F7 to n‐C4F9.
Longer tails are more attractive: Electron affinities of aromatic compounds demonstrate that the RF substituent electron‐withdrawing ability increases as CF3<C2F5<n‐C3F7<n‐C4F9.
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