High-precision mapping reveals rare N6-deoxyadenosine methylation in the mammalian genome.

N6-deoxyadenosine methylation (6mA) is the most widespread type of DNA modification in prokaryotes and is also abundantly distributed in some unicellular eukaryotes. However, 6mA levels are remarkably low in mammals. The lack of a precise and comprehensive mapping method has hindered more advanced investigations of 6mA. Here, we report a new method MM-seq (modification-induced mismatch sequencing) for genome-wide 6mA mapping based on a novel detection principle. We found that modified DNA bases are prone to form a local open region that allows capture by antibody, for example, via a DNA breathing or base-flipping mechanism. Specified endonuclease or exonuclease can recognize the antibody-stabilized mismatch-like structure and mark the exact modified sites for sequencing readout. Using this method, we examined the genomic positions of 6mA in bacteria (E. coli), green algae (C. reinhardtii), and mammalian cells (HEK239T, Huh7, and HeLa cells). In contrast to bacteria and green algae, human cells possess a very limited number of 6mA sites which are sporadically distributed across the genome of different cell types. After knocking out the RNA m6A methyltransferase METTL3 in mouse ES cells, 6mA becomes mostly diminished. Our results imply that rare 6mA in the mammalian genome is introduced by RNA m6A machinery via a non-targeted mechanism.

[The spectrogram characteristics of organic blue-emissive light-emitting excitated YAG : Ce phosphor].

It is demonstrated that the panchromatic luminescence devices with organic blue-emissive light-emitting was fabricated. This technique used down conversion, which was already popular in inorganic power LEDs to obtain white light emission. A blue OLED device with a configuration of ITO/2T-NATA (30 nm)/AND : TBPe (50 Wt%, 40 nm)/Alq3 (100 nm)/LiF(1 nm)/Al(100 nm) was prepared via vacuum deposition process, and then coated with YAG : Ce phosphor layers of different thicknesses to obtain a controllable and uniform shape while the CIE coordinates were fine tuned. This development not only decreased steps of technics and degree of difficulty, but also applied the mature technology of phosphor. The results showed that steady spectrogram was obtained in the devices with phosphor, with a best performance of a maximum luminance of 13 840 cd x m(-2) which was about 2 times of that of the devices without phosphor; a maximum current efficiency of 17.3 cd x A(-1) was increased more two times more than the devices without phosphor. The emission spectrum could be adjusted by varying the concentration and thickness of the phosphor layers. Absoulte spectrogram of devices was in direct proportion with different driving current corresponding.

Pharmacology of the urotensin-II receptor antagonist palosuran (ACT-058362; 1-[2-(4-benzyl-4-hydroxy-piperidin-1-yl)-ethyl]-3-(2-methyl-quinolin-4-yl)-urea sulfate salt): first demonstration of a pathophysiological role of the urotensin System.

Urotensin-II (U-II) is a cyclic peptide now described as the most potent vasoconstrictor known. U-II binds to a specific G protein-coupled receptor, formerly the orphan receptor GPR14, now renamed urotensin receptor (UT receptor), and present in mammalian species. Palosuran (ACT-058362; 1-[2-(4-benzyl-4-hydroxy-piperidin-1-yl)-ethyl]-3-(2-methyl-quinolin-4-yl)-urea sulfate salt) is a new potent and specific antagonist of the human UT receptor. ACT-058362 antagonizes the specific binding of (125)I-labeled U-II on natural and recombinant cells carrying the human UT receptor with a high affinity in the low nanomolar range and a competitive mode of antagonism, revealed only with prolonged incubation times. ACT-058362 also inhibits U-II-induced calcium mobilization and mitogen-activated protein kinase phosphorylation. The binding inhibitory potency of ACT-058362 is more than 100-fold less on the rat than on the human UT receptor, which is reflected in a pD'(2) value of 5.2 for inhibiting contraction of isolated rat aortic rings induced by U-II. In functional assays of short incubation times, ACT-058362 behaves as an apparent noncompetitive inhibitor. In vivo, intravenous ACT-058362 prevents the no-reflow phenomenon, which follows renal artery clamping in rats, without decreasing blood pressure and prevents the subsequent development of acute renal failure and the histological consequences of ischemia. In conclusion, the in vivo efficacy of the specific UT receptor antagonist ACT-058362 reveals a role of endogenous U-II in renal ischemia. As a selective renal vasodilator, ACT-058362 may be effective in other renal diseases.

Chronic endothelin receptor blockade prevents both early hyperfiltration and late overt diabetic nephropathy in the rat.

Diabetic nephropathy is associated with enhanced renal synthesis of endothelin (ET)-1. The goal of this study was to investigate the effects of dual ET receptor antagonism in the early phase (2 months) and in the late phase (5 months) of diabetic nephropathy in rats, and to compare this approach to angiotensin-converting enzyme inhibition. Four groups of uninephrectomized streptozotocin-induced diabetic rats were assigned to receive orally vehicle, bosentan, enalapril, or their combination. A fifth group consisted of nondiabetic, uninephrectomized rats. At 2 weeks, untreated diabetic rats exhibited increased glomerular filtration rate and renal plasma flow. Bosentan, enalapril, and the combination all prevented hyperfiltration and hyperperfusion. By 5 months, diabetic rats developed marked increases in mean arterial pressure and renal vascular resistance, progressive proteinuria, and renal structural damage with glomerular sclerosis and hypertrophy. Bosentan completely prevented the development of hypertension and renal vasoconstriction, and largely prevented the development of proteinuria and renal structural injury. The renal protective effect of bosentan was comparable to that of enalapril or the combination, although its anti-proteinuric effect was less. Clinical studies are warranted to assess whether ET receptor antagonism can have additive effects on top of ACE inhibition, the current treatment of choice in diabetic nephropathy.

Chronic endothelin receptor blockade prevents renal vasoconstriction and sodium retention in rats with chronic heart failure.

OBJECTIVE: Importance of endothelin in mediating the chronic renal alterations of chronic heart failure was studied in rats chronically treated with bosentan after myocardial infarction. METHODS: Rats were subjected to coronary artery ligation and were treated for 8 weeks with placebo or bosentan, a dual ET(A) and ET(B) receptor antagonist, (approximately 100 mg/kg/day) as food admix. Sham-operated rats served as normal controls. Cardiac and renal functions were measured at the end of 8-week treatment. RESULTS: Bosentan significantly reduced the elevated left ventricular end-diastolic pressure (from 26.6+/-3.3 to 11.4+/-2.2 mmHg, P<0.001) and the increased heart-to-body-weight ratio seen in untreated rats with myocardial infarction. Bosentan prevented the marked increase in renal vascular resistance (bosentan, 7.7+/-0.6; untreated, 15.6+/-2.5 mmHg/ml/min; P<0.001). This led to a significant increase in renal plasma flow resulting in a decrease in filtration fraction. Bosentan furthermore increased urinary sodium excretion. CONCLUSIONS: Prolonged ET receptor blockade in rats with myocardial infarction has chronic renal vasodilatory effect and improves renal sodium excretory function. Thus, dual ET antagonists such as bosentan might be useful in the treatment of the progressive renal failure associated with human chronic heart failure.