Hydrolytic reactions of the phosphorodithioate analogue of uridylyl(3',5')uridine: kinetics and mechanisms for the cleavage, desulfurization, and isomerization of the internucleosidic linkage

The hydrolytic reactions of the phosphorodithioate analogue of uridylyl(3',5')uridine [3',5'-Up(s)2U] were followed by HPLC over a wide pH range at 363.2 K. Under acidic and neutral conditions, three reactions compete: (i) desulfurization to a mixture of the (Rp)- and (Sp)-diastereomers of the corresponding 3',5'- and 2',5'-phosphoromonothioates [3',5'- and 2',5'-Up(s)U], which are subsequently desulfurized to a mixture of uridylyl(3',5')- and -(2',5')uridine [3',5'- and 2',5'-UpU], (ii) isomerization to 2',5'-Up(s)2U, and (iii) cleavage to uridine, in all likelihood via a 2',3'-cyclic phosphorodithioate (2',3'-cUMPS2). Under alkaline conditions (pH > 8), only a hydroxide ion catalyzed hydrolysis to uridine via 2',3'-cUMPS2 takes place. At pH 3-7, all three reactions are pH-independent, the desulfurization being approximately 1 order of magnitude faster than the cleavage and isomerization. At pH < 3, all the reactions are hydronium ion catalyzed. On going to very acidic solutions, the cleavage gradually takes over the desulfurization and isomerization. Accordingly, the cleavage overwhelmingly predominates at pH < 0. The overall hydrolytic stability of 3',5'-Up(s)2U is comparable to that of (Sp)- and (Rp)-3',5'-Up(s)U (and to that of 3',5'-UpU, except at pH < 2). The rate of the hydroxide ion catalyzed hydrolysis of 3',5'-Up(s)2U is 37% and 53% of that of (Sp)- and (Rp)-3',5'-Up(s)U, respectively. The reactions, however, differ with the respect of the product accumulation. While the phosphoromonothioates produce a mixture of 2'- and 3'-thiophosphates as stable products, 3',5'-Up(s)2U is hydrolyzed to uridine without accumulation of the corresponding dithiophosphates. At pH < 3, where the hydrolysis is hydronium ion catalyzed, the kinetic thio-effect of the second thio substitution is small: under very acidic conditions (Ho -0.69), (Sp)-3',5'-Up(s)U reacts 1.6 times as fast as 3',5'-Up(s)2U, but the reactivity difference decreases on going to less acidic solutions. In summary, the hydrolytic stability of 3',5'-Up(s)2U closely resembles that of the corresponding phosphoromonothioate. While replacing one of the nonbridging phosphate oxygens of 3',5'-UpU with sulfur stabilizes the phosphodiester bond under acidic conditions by more than 1 order of magnitude, the replacement of the remaining nonbridging oxygen has only a minor influence on the overall hydrolytic stability.

Detection of high-frequency respiratory movements during sleep.

Sleep-related breathing disorders are common in adults and they have a significant impact on vigilance and quality of life. Previous studies have shown the validity of the static-charge-sensitive bed (SCSB) in monitoring breathing abnormalities during sleep. A whole nights sleep study produces a signal with considerable length, and therefore an automated analysis system would be of great need. In this work we focus on detection of high-frequency respiratory movement (HFRM) patterns which are related to increased respiratory efforts. The paper documents four methods to automatically detect these patterns. The first two are based on classical statistical tests applied to the SCSB signal, and the other two use spectral characteristics in order to adaptively segment the SCSB signal. Finally we adjust each method to detect patterns that coincide with the HFRMs determined by an expert, and evaluate the performance of the methods using independent test data.

Choroidal readaptation to gravity in rats after spaceflight and head-down tilt.

To determine when choroidal structures were restored after readaptation to Earth gravity or orthostatic position, fine structure and protein distribution were studied in rat choroid plexus dissected either 6 h [Space Life Sciences-2 (SLS-2) experiments] or 2 days [National Institutes of Health-Rodent 1 (NIH-R1) experiments] after a spaceflight, or 6 h after head-down tilt (HDT) experiments. Apical alterations were noted in choroidal cells from SLS-2 and HDT animals, confirming that weightlessness impaired choroidal structures and functions. However, the presence of small apical microvilli and kinocilia and the absence of vesicle accumulations showed that the apical organization began to be restored rapidly after landing. Very enlarged apical microvilli appeared after 2 days on Earth, suggesting increased choroidal activity. However, as distributions of ezrin and carbonic anhydrase II remained altered in both flight and suspended animals after readaptation to Earth gravity, it was concluded that choroidal structures and functions were not completely restored, even after 2 days in Earth's gravity.

A PC program for automatic analysis of NMR spectrum series.

31P nuclear magnetic resonance (NMR) spectroscopy enables us to study intracellular energy metabolism noninvasively. The present work concerns the analysis of a series of 31P NMR spectra on human muscle during exercise. The spectra contain signals corresponding to certain metabolites in the sample and the aim is to identify and quantify these signals. We have written a PC program to perform this task automatically. With the program the results can be achieved substantially faster compared to operating with the spectrometer software. The methods implemented in the program and the functions of the program itself are described. Although we have focused on the 31P NMR spectrum series, the program can also be applied to other liquid state NMR spectra.

Prostacyclin and thromboxane in chronic uremia: effect of hemodialysis.

To study the effects of uremia and hemodialysis on the production rates of antiaggregatory prostacyclin (PGI2) and proaggregatory thromboxane A2 (TxA2), we collected serial plasma samples from eight patients with chronic uremia before, during and after hemodialysis and assayed them for 6-keto-PGF1 alpha and TxB2, the stable metabolites of PGI2 and TxA2, respectively. In addition, the capacity of the platelets to produce TxB2 during spontaneous clotting was studied by measuring the TxB2 levels in serum incubated at +37 degrees C for 60 minutes. The PGI2 production of the uremia patients before hemodialysis was less (P less than 0.001) than that of healthy volunteers. It rose significantly following heparinization and remained elevated during hemodialysis. TxB2 generation by platelets during clotting was diminished in uremia. Plasma TxB2 levels were normal before, but increased during hemodialysis. Thus, profound changes in the PGI2/TxA2-system seem to be associated with uremia and hemodialysis.

Nephropathia epidemica. The Scandinavian form of hemorrhagic fever with renal syndrome.

We report epidemiologic, clinical, laboratory, and biopsy findings in 14 cases of nephropathia epidemica. The patients were between 19 and 49 years of age. The onset of the disease was characterized by high fever, nausea, headache, backache, abdominal pain, proteinuria, oliguria, hematuria, and uremia. The symptoms subsided rapidly during the polyuria phase, which followed the oliguria stage. Because of renal failure, hemodialysis was required in eight cases. Edema of eyelids, conjunctival injection and hemorrhages, transitory myopia, and acute glaucoma were the most common eye abnormalities. Renal biopsy specimens showed glomerular changes, with mild swelling of the epithelial cells of Bowman's capsule, thickening of the basement membrane of glomerular capillaries, glomerular adhesions, inflammatory cell infiltration, leukocytoclasis and hemorrhages in the interstitium, and eosinophilic hyaline degeneration and vacuolization of the epithelial cells of the proximal tubuli.

Triglycerides, cholesterol, and phospholipids in normal heart papillary muscle and in patients suffering from diabetes, cholelithiasis, hypertension, and coronary atheroma.

The triglyceride, cholesterol, and phospholipid contents of heart papillary muscle were measured in groups of obviously healthy and diseased females and males on whom either routine or forensic necropsies were performed. In healthy men the triglyceride content was 1.77 +/- 1.30 mg/g of wet weight and in women 1.25 +/- 0.48 mg/g wet weight. The corresponding values for cholesterol were 1.07 +/- 0.24 mg/g and 1.21 +/- 0.22 mg/g and those for phospholipids 17.70 +/- 5.15 mg/g and 19.65 +/- 10.21 mg/g. The differences between the sexes were not significant. The hypertensive or cardiac hypertrophy group had about the same or slightly lower means for lipid content. In the cholelithiasis group, women had significantly high triglyceride values (3.38 +/- 2.36 mg/g). The cholesterol values were not significantly elevated in either men or women. In the diabetic group, triglycerides were significantly increased both in men (mean 8.12 +/- 0.54 mg/g) and in women (6.85 +/- 5.66 mg/g). The cholesterol mean values were also high in both sexes, but the rise was not significant because of the great variation. In the coronary atheroma group, both male and female hospital cases had high triglyceride contents (mean 4.48 +/- 4.25 mg/g and 3.65 +/- 3.94 mg/g) whereas the forensic cases had only slightly elevated or normal values. Cholesterol assays paralleled the triglyceride ones, but phospholipids showed an inverse trend. The results showed that the lipid content of papillary muscle was increased in diseases where disturbances of lipid metabolism are evident, as in diabetes and cholelithiasis. In coronary atheroma only those cases with advanced obstruction of the arteries were associated with abnormal values of papillary lipids. No increase of the lipid content with age alone was found, nor was there any correlation with obesity.