It is unclear whether thiazide diuretics (TZs) or calcium channel blockers (CCBs) are more effective as add-on therapy to angiotensin receptor blockers (ARBs) in controlling hypertension. Because TZs ...are a rational choice in salt-sensitive hypertension, patients with high salt intake might preferentially benefit from ARB/TZ over ARB/CCB combination therapy.
Hypertensive patients who failed to reach blood pressure goals despite treatment with ARBs alone were randomly assigned to receive either ARB/TZ or ARB/CCB combination therapy. Estimated daily sodium intake was calculated from spot urine values of sodium and creatinine.
Blood pressure was measured at baseline, and at 4, 8 and 12 weeks after starting combination therapy. For all study patients (n = 87), diastolic blood pressure reduction was greater in patients receiving ARB/CCB treatment. However, in the 37 patients with a baseline estimated daily salt intake greater than 10 g and baseline systolic blood pressure (SBP) ranging from 150 to 200 mm Hg, SBP was lower (P < 0.05) and SBP reduction was greater (P < 0.05) 4 weeks after starting combination therapy in those receiving ARB/TZ treatment. In the 31 patients whose estimated daily salt intake increased at 12 weeks compared with baseline, SBP at 12 weeks was lower in those receiving ARB/TZ treatment (P < 0.05).
Estimated daily salt intake is a useful tool for guiding antihypertensive therapy and should be measured repeatedly during the therapeutic course.
DNA-dependent protein kinase (DNA-PK) is composed of a 460-kDa catalytic component (p460) and a DNA-binding component Ku protein. Immunoblot analysis after treatment of Jurkat cells with anti-Fas ...antibody demonstrated the cleavage of p460 concomitantly with an increase in CPP32/Yama/apopain activity. Recombinant CPP32/Yama/apopain specifically cleaved p460 in the DNA-PK preparation that had been purified from Raji cells into 230- and 160-kDa polypeptides, the latter of which was detected in anti-Fas-treated Jurkat cells. The regulatory component Ku protein was not significantly affected by CPP32/Yama/apopain. DNA-PK activity was decreased with the disappearance of p460 in the incubation of DNA-PK with CPP32/Yama/apopain. These results suggest that the catalytic component of DNA-PK is one of the target proteins for CPP32/Yama/apopain in Fas-mediated apoptosis.
A double-stranded DNA-dependent protein serine/threonine kinase (DNA-PK) was purified from a nuclear extract of Raji Burkitt's lymphoma cells by a three-step column-chromatographic procedure. The ...main silver-stained band visualized after SDS/PAGE corresponded to an autophosphorylated polypeptide of about 350-kDa that represents the catalytic component. The existence of Ku DNA-binding protein as a regulatory component in the purified enzyme was revealed by Western blot/enzyme immunoassay and direct inhibition test with anti-Ku sera from the autoimmune patients. The DNA-PK catalyzed phosphorylation of synthetic peptides corresponding to Myc and RB proteins in a DNA-dependent manner, indicating that DNA-PK may recognize a second core-sequence motif Pro-Ser/Thr- in addition to the putative consensus sequences of -Ser/Thr-Gln. The level of enzyme activity was significantly higher in DMSO-induced G0/G1-arrested Raji cells as well as in the cells after release from DMSO than in the log-phase cells.
DNA-dependent protein kinase (DNA-PK), consisting of the 470-kDa catalytic component (DNA-PKcs) and the DNA-binding regulatory component Ku protein (p70/p80), catalyzes phosphorylation of a variety ...of DNA replication/transcription/repair factors in the presence of double-stranded DNA. In the resting states of human peripheral blood mononuclear cells, DNA-PK activity and the protein level of DNA-PKcs were very low in the nuclear extracts, but they were high in the whole cell extracts. Depending upon proliferation of the T lymphocytes, DNA-PK activity and the protein level of DNA-PKcs in the nuclear extracts greatly increased. Immunocytochemical analysis suggested translocation of DNA-PKcs from the cytoplasm to the nucleus upon growth stimulation in the T lymphocytes.
S-Adenosylmethionine synthetase exists in at least two distinct forms, α- and β-forms, in adult liver. The β-form was purified to homogeneity from the soluble fraction of rat liver with a yield of ...about 10%. An antiserum directed against the purified β-form from rat liver was prepared by injecting the purified enzyme into a rabbit. Ouchterlony double diffusion analysis and immunochemical titrα-tions revealed that the isozymes, α- and β-forms, are identical. Thus, the α-form was isolated from rat liver as a single protein using immunoaffinity chromatography against the β-form. The molecular weights of the β- and α-forms were determined to be 48,000 each by sodium dodecyl sulfate disc gel electrophoresis, and about 100,000, and 200,000, respectively, by Sephacryl S-200 gel fitration. These results indicate that the β-form consisted of two subunits of 48,000 daltons and the α-form of four subunits of 48,000 daltons. The sedimentation coefficient was calculated to be 5.5S for the β-form and 8.0S for the α-form.
cDNA clones encoding the human kidney
S-adenosylmethionine synthetase (kidney-type isozyme) were isolated. The amino acid sequence deduced from the cDNA indicates that this enzyme contains 395 amino ...acids and has a molecular mass of 43,660 Da. The predicted amino acid sequence of this protein shares 84% similarity with that of human liver
S-adenosylmethionine synthetase (liver-type isozyme). In addition, the developmental expression of these two isozyme mRNAs has been studied in the human liver using the reverse transcription—polymerase chain reaction (RT-PCR).
DNA‐dependent protein kinase (DNA‐PK) has been known to catalyze phosphorylation of a number of regulatory factors involved in DNA replication and transcription such as simian virus 40 T antigen, ...p53, c‐Myc, Spl, and RNA polymerase II (Pol II). We examined the possibility that DNA‐PK phosphory‐lates the general transcription factors TATA‐binding protein (TBP) and transcription factor (TF) IIB, which play key roles in the formation of transcription initiation complex with Pol II. By using a highly purified preparation of DNA‐PK from Raji cells, both TBP and TFIIB were shown to be phosphorylated in vitro by DNA‐PK. We then investigated the effect of the phosphorylation of these factors on Pol II basal transcription. Stepwise analysis of preinitiation complex formation by electrophoretic mobility shift assay revealed that the phosphorylation of TBP and TFIIB by DNA‐PK did not affect the formation of promoter (P)‐TBP and P‐TBP‐TFIIB complexes but synergistically stimulated the formation of P‐TBP‐TFIIB‐TFIIF‐Pol II complex. Similarly, combination of the phosphorylated TBP and TFIIB synergistically stimulated Pol II basal transcription from adenovirus major late promoter. These observations suggest that DNA‐PK could positively regulate the Pol II basal transcription by phosphorylating TBP and TFIIB.
The activities of S-adenosylmethionine synthetase isozymes in liver were measured after rats received a diet containing excess methionine. The activity of the α-form increased with increasing ...methionine content in the diet, and reached 4–5 fold after 6 days on a 3 % methionine diet. However, the activity of the β-form showed only a 1.5 fold increase. The activity of the γ-form in kidney showed no significance change.
Two RNases H, Mg2+- and Mn2+-dependent RNases H, are present in extracts of chick embryo. These RNases H can be separated by phosphocellulose column chromatography. Mg2+-dependent RNase H was ...purified over 900-fold and Mn2+-dependent RNase H over 1,700-fold from chick embryo extracts. The molecular weight of the purified Mg2+-dependent RNase H was about 40,000 and of the Mn2+-dependent RNase H about 120,000, when estimated by gel filtration. Mg2+-dependent RNase H exhibits maximal activity at pH 9.5, and requires 15 to 20 mM Mg2+ for maximal activity, whereas Mn2+-dependent RNase H is most active at pH 8.5, and is maximally active at the concentration of 0.4 mM Mn2+, and has some activity with Mg2+. Both enzymes require a sulfhydryl reagent for maximal activity. Mn2+-dependent RNase H was inhibited by o-phenanthroline, pyrophosphate, and those polyamines tested, whereas Mg2+-dependent enzyme was not, although it was inhibited by NaF. Both RNases H liberate a mixture of oligonucleotides with 5′-phosphate and 3′-hydroxyl termini endonucleolytically.
Treatment of rats with an ethionine plus adenine or a methionine diet leads not only to a marked increase of the α-form isozyme of S-adenosylmethionine synthetase in liver, but also to the ...accumulation of comparable amounts of S-adenosylethionine and S-adenosylmethionine in liver. Transplantation of ascites tumor cells into mice leads to a marked increase only of the β-form isozyme in the host liver, but the levels of S-adenosylmethionine do not significantly change in liver.
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