Sex Difference in Plasma Deoxyribonuclease Activity in Rats.

Extracellular DNA (ecDNA) activates immune cells and is involved in the pathogenesis of diseases associated with inflammation such as sepsis, rheumatoid arthritis or metabolic syndrome. DNA can be cleaved by deoxyribonucleases (DNases), some of which are secreted out of cells. The aim of this experiment was to describe plasma DNase activity in relation to extracellular DNA in adult rats, to analyse potential sex differences and to prove whether they are related to endogenous testosterone. Adult Lewis rats (n=28) of both sexes were included in the experiment. Male rats were gonadectomized or sham-operated and compared to intact female rats. Plasma ecDNA and DNase activity were measured using fluorometry and single radial enzyme diffusion assay, respectively. Concentrations of nuclear ecDNA and mitochondrial ecDNA were determined using real-time PCR. Females had 60% higher plasma DNase activity than males ( p=0.03). Gonadectomy did not affect plasma DNase in males. Neither the concentration of total ecDNA, nor nuclear or mitochondrial DNA in plasma differed between the groups. No significant correlations between DNase and ecDNA were found. From previous studies on mice, it was expected, that male rats will have higher DNase activity. In contrast, our study in rats showed the opposite sex difference. This sex difference seems not to be caused by endogenous testosterone. Interestingly, no sex differences were observed in plasma ecDNA suggesting a complex or missing association between plasma ecDNA and DNase. The observed sex difference in plasma DNase should be taken into account in animal models of ecDNA-associated diseases.

Does the 2nd and 4th digit ratio reflect prenatal androgen exposure?

OBJECTIVES:  The aim of our study was to describe the effect of prenatal testosterone exposure on 2D:4D in both sexes, and to determine whether this effect is mediated via the androgen receptor. In addition, the sex differences in lengths of 2D, 4D, and 2D:4D ratio were analyzed. BACKGROUND:  Clinical studies suggest a negative correlation between prenatal testosterone exposure and ratio of the lengths of the second and fourth digits (2D:4D). However, less is known about the underlying molecular mechanisms. METHODS:  Pregnant rats were treated with olive oil, testosterone, flutamide or testosterone with flutamide daily from the fourteenth day of pregnancy until delivery. The finger lengths of adult offspring were measured using both, digital scanning of the paws and µCT analysis of the phalanges. RESULTS:  None of the aforementioned methods revealed any effect of testosterone on 2D:4D. µCT measurements showed that prenatal hyperandrogenism in both sexes leads to shorter 2D compared to controls. Moreover, the testosterone treatment in males resulted in the shortening of 4D when compared to controls. CONCLUSION:  Prenatal hyperandrogenism leads to shorter lengths of 2D and 4D; however, it does not affect 2D:4D ratio. Whether other steroid hormones and/or testosterone metabolites affect the 2D:4D ratio requires further investigation (Tab. 5, Fig. 3, Ref. 32).

Extracellular DNA as a Prognostic and Therapeutic Target in Mouse Colitis under DNase I Treatment.

The aim of this study was to investigate the potential of extracellular DNA as a prognostic and/or therapeutic target in inflammatory bowel disease. Fifty male C57BL/6J mice were used in the experiment. Acute colitis was induced by intake of 2% dextran sulphate sodium (DSS) for seven days followed by three days of water intake. DNase I was injected intravenously on days 3 and 7. Plasmatic levels of extracellular DNA (ecDNA) were measured on days 6 and 10. Weight loss, stool consistency and liquid intake were monitored throughout the experiment. Colon length and weight, myeloperoxidase activity and tumour necrosis factor α (TNF-α) levels were measured at sacrifice. DSS-treated mice displayed severe colitis, as shown by disease activity parameters. Both groups with colitis (DNase treated and untreated) had significantly poorer weight loss, colon length and stool consistency compared with control groups on water. No differences between the DNasetreated and untreated DSS groups were recorded. Myeloperoxidase activity and levels of TNF-α in colonic tissue were notably greater in both groups with colitis compared to controls. In addition, both biochemical markers were improved in the DNasetreated group with colitis compared to the untreated group. Although the disease activity was proved by several independent parameters in both groups with colitis, levels of ecDNA did not show any difference between the groups throughout or at the end of experiment. The role of ecDNA in experimental colitis has not been confirmed. However, DNase I injection resulted in some improvement, and thus should be studied in more detail.

Immune activation by nucleic acids: A role in pregnancy complications.

Cell-free self-DNA or RNA may induce an immune response by activating specific sensing receptors. During pregnancy, placental nucleic acids present in the maternal circulation further activate these receptors due to the presence of unmethylated CpG islands. A higher concentration of cell-free foetal DNA is associated with pregnancy complications and a higher risk for foetal rejection. Cell-free foetal DNA originates from placental trophoblasts. It appears in different forms: free, bound to histones in nucleosomes, in neutrophil extracellular traps (NETs) and in extracellular vesicles (EVs). In several pregnancy complications, cell-free foetal DNA triggers the production of proinflammatory cytokines, and this production results in a cellular and humoral immune response. This review discusses preeclampsia, systemic lupus erythematosus, foetal growth restriction, gestational diabetes, rheumatoid arthritis and obesity in pregnancy from an immunological point of view and closely examines the different pathways that result in maternal inflammation. Understanding the role of cell-free nucleic acids, as well as the biogenesis of NETs and EVs, will help us to specify their functions or targets, which seem to be important in pregnancy complications. It is still not clear whether higher concentrations of cell-free nucleic acids in the maternal circulation are the cause or consequence of various complications. Therefore, further clinical studies and, even more importantly, animal experiments that focus on the involved immunological pathways are needed.

Does rat fetal DNA induce preeclampsia in pregnant rats?

Cell-free fetal DNA in maternal circulation is higher during preeclampsia. It is unclear whether it is the cause or the consequence of the disease. The aim of this study was to prove whether injected rat fetal DNA induces preeclampsia-like symptoms in pregnant Wistar rats. They received daily i.p. injections of water or rat fetal DNA (400 µg) from gestation day 14 to 18. Blood pressure, proteinuria, placental and fetal weight were measured at gestation day 19. Plasma DNase activity, proteinuria and creatinine clearance were assessed. There was no significant difference in any of the measured parameters. The results of this study do not confirm the hypothesis that fetal DNA might induce preeclampsia. This is in contrast to others using human fetal DNA in mice. Further studies should be focused on the effects of fetal DNA from the same species protected from DNase activity.

Origin of choriocarcinoma in previous molar pregnancy proved by DNA analysis.

A 17-year old woman had in a short time period (seven months) a very exciting reproduction history. Molar pregnancy in December 1993, choriocarcinoma in January 1994 and induced abortion in June 1994. DNA analysis proved the origin of the choriocarcinoma in the previous molar pregnancy.