Cell-free DNA in medium is associated with the maturation ability of in vitro cultured oocytes.

Follicular fluid contains cell-free DNA (cfDNA), which may serve as a useful biomarker of oocyte ability. The present study evaluates whether nuclear and mitochondrial cfDNAs in conditioned oocyte growth medium determine the quality of oocytes cultured in vitro. Oocyte and granulosa cell complexes (OGCs) derived from early antral follicles of gilt ovaries were cultured for 14 days and the amount of cfDNA and lactate concentration in the conditioned culture medium were measured and compared to evaluate oocyte maturation ability. The amount of nuclear cfDNA, but not mitochondrial cfDNA, strongly correlated with the number of dead cells in OGCs. Furthermore, low mitochondrial cfDNA content and high lactate concentration in the medium was associated with high maturation ability of oocytes cultured in vitro. In conclusion, the amounts of nuclear and mitochondrial cfDNAs differentially reflect the conditions of OGCs, and low mitochondrial cfDNA, low glucose content, and high lactate concentration in the medium are associated with the proper maturation of oocytes.

Elevated Plasma Levels of Cell-Free DNA During Liver Transplantation Are Associated With Activation of Coagulation.

Patients undergoing liver transplantation have complex changes in their hemostatic system, and the net effect of these changes appears to be a "rebalanced" hemostatic profile. Recently, a process called NETosis in which a neutrophil expels DNA and proteins that form a weblike structure, has been described as a mechanism of pathogen entrapment. Increasing evidence suggests a pivotal role for neutrophil extracellular traps (NETs) and their main component, cell-free DNA (cfDNA), in activation of coagulation. Because liver transplantation is associated with substantial (hepatocyte) cell death and intrahepatic neutrophil accumulation, NETs might play an important role in the hemostatic balance during liver transplantation. Here, we determined markers for NETs in the plasma of patients undergoing a liver transplantation and examined their association with activation of coagulation. Markers for NETs and markers for activation of coagulation were determined in serial plasma samples taken from patients undergoing a liver transplantation (n = 21) and compared with plasma levels in healthy controls. We found perioperative increases of markers for NETs with levels of cfDNA and nucleosomes that peaked after reperfusion and myeloperoxidase (MPO)-DNA complexes that peaked during the anhepatic phase. CfDNA and nucleosome levels, but not MPO-DNA levels, correlated with prothrombin fragment 1+2 and thrombin-antithrombin complex levels, which are established markers for activation of coagulation. Neutrophils undergoing NETosis were observed by immunostainings in postreperfusion biopsies. In conclusion, although NETosis occurs during liver transplantation, the majority of circulating DNA appears to be derived from cell death within the graft. The perioperative increases in cfDNA and nucleosomes might contribute to the complex hemostatic rebalance during liver transplantation.

Extracellular self-DNA as a damage-associated molecular pattern (DAMP) that triggers self-specific immunity induction in plants.

Mammals sense self or non-self extracellular or extranuclear DNA fragments (hereinafter collectively termed eDNA) as indicators of injury or infection and respond with immunity. We hypothesised that eDNA acts as a damage-associated molecular pattern (DAMP) also in plants and that it contributes to self versus non-self discrimination. Treating plants and suspension-cultured cells of common bean (Phaseolus vulgaris) with fragmented self eDNA (obtained from other plants of the same species) induced early, immunity-related signalling responses such as H2O2 generation and MAPK activation, decreased the infection by a bacterial pathogen (Pseudomonas syringae) and increased an indirect defence to herbivores (extrafloral nectar secretion). By contrast, non-self DNA (obtained from lima bean, Phaseolus lunatus, and Acacia farnesiana) had significantly lower or no detectable effects. Only fragments below a size of 700 bp were active, and treating the eDNA preparation DNAse abolished its inducing effects, whereas treatment with RNAse or proteinase had no detectable effect. These findings indicate that DNA fragments, rather than small RNAs, single nucleotides or proteins, accounted for the observed effects. We suggest that eDNA functions a DAMP in plants and that plants discriminate self from non-self at a species-specific level. The immune systems of plants and mammals share multiple central elements, but further work will be required to understand the mechanisms and the selective benefits of an immunity response that is triggered by eDNA in a species-specific manner.

The role of total cell-free DNA in predicting outcomes among trauma patients in the intensive care unit: a systematic review.

BACKGROUND: Cell-free DNA has been proposed as a means of predicting complications among severely injured patients. The purpose of this systematic review was to assess whether cell-free DNA was useful as a prognostic biomarker for outcomes in trauma patients in the intensive care unit. METHODS: We searched Pubmed, Embase, Scopus and the Cochrane Central Register for Controlled Trials and reference lists of relevant articles for studies that assessed the prognostic value of cell-free DNA detection in trauma patients in the intensive care unit. Outcomes of interest included survival, posttraumatic complications and severity of trauma. Due to considerable heterogeneity between the included studies, a checklist was formed to assess quality of cell-free DNA measurement. RESULTS: A total of 14 observational studies, including 904 patients, were eligible for analysis. Ten studies were designed as prospective cohort studies; three studies included selected patients from a cohort while one study was of a retrospective design. We found a significant correlation between higher values of cell-free DNA and higher mortality. This significant correlation was evident as early as on intensive care unit admission. Likewise, cell-free DNA predicted the severity of trauma and posttraumatic complications in a majority of patients. CONCLUSION: The amount of cell-free DNA can function as a prognostic tool for mortality and to a lesser extent severity of trauma and posttraumatic complications. Standardizing cell-free DNA measurement is paramount to ensure further research in cell-free DNA as a prognostic tool.

The roles of cirRNA in the development of germ cells.

Circular RNA (CircRNA), a type of endogenous non-coding RNAs (ncRNAs), is generally generated from precursor mRNA (pre-mRNA) by canonical splicing and head-to-tail back splicing. The structure without a polyA tail renders circRNA highly insensitive to ribonuclease. Simultaneously, the distribution of circRNAs is tissue and developmental stage-specific. There are five potential biological functions of circRNAs: 1) promote transcription of their parental genes; 2) function as a miRNA sponge; 3) RNA binding protein (RBP) sponge; 4) encode protein; 5) act as an mRNA trap. Recently, circRNA has attracted attention because studies have shown that circRNAs are associated with follicular development, ovarian senescence, spermatogenesis, and germ cell development process, suggesting that circRNAs may function in germ cells regulation. The investigation of circRNAs in germ cells will provide an excellent opportunity to understand its potential molecular basis, and potentially improving reproduction status in human. In this article, the relationship between circRNA and germ cell development will be discussed.

Inflammation-mediated generation and inflammatory potential of human placental cell-free fetal DNA.

INTRODUCTION: Circulating DNA can be pro-inflammatory when detected by leukocytes via toll-like receptor 9 (TLR9). Cell-free fetal DNA (cff-DNA) of placental origin, circulates in pregnancy, and increased concentrations are seen in conditions associated with placental and maternal inflammation such as pre-eclampsia. However, whether cff-DNA is directly pro-inflammatory in pregnant women and what regulates cff-DNA levels in pregnancy are unknown. METHODS: Using a human term placental explant model, we examined whether induction of placental inflammation can promote cff-DNA release, and the capacity of this cff-DNA to stimulate peripheral blood mononuclear cells (PBMCs) from pregnant women. RESULTS: We demonstrate lipopolysaccharide (LPS)-mediated inflammation in placental explants and induced apoptosis after 24 h. However, this did not increase levels of cff-DNA generation compared to controls. Furthermore, the methylation status of the cff-DNA, was not altered by LPS-induced inflammation. Cff-DNA did not elicit production of inflammatory cytokines from PBMCs, in contrast to exposure to LPS or the TLR9 agonist CpG-ODN. Finally, we demonstrate that cff-DNA acquired directly from pregnant women did not differ in methylation status from placental extracted DNA, or from placental explant generated cell-free DNA, and that, unlike Escherichia coli DNA, this cff-DNA has a low level of unmethylated CpG sequences. DISCUSSION: Our data suggest that placental inflammation does not increase release of cff-DNA and that placental cff-DNA is not pro-inflammatory to circulating PBMCs. It thus seems unlikely that high levels of cff-DNA are either a direct consequence or cause of inflammation observed in obstetric complications.