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J P Rieker

Publications and source records attributed to J P Rieker.

8 recordsLinked to original sources

Teaching critical appraisal during critical care fellowship training: a foundation for evidence-based critical care medicine.

OBJECTIVE: To determine whether fellowship training in critical care medicine with critical appraisal exercises improves the ability and confidence of fellows to evaluate the medical literature. DESIGN: Prospective, interventional pilot study. SETTING: Multidisciplinary critical care medicine training program at a large university hospital. INTERVENTION: Fellows were given three didactic sessions covering study design, analysis, and critical appraisal techniques. During the course of the year, each fellow was required to review one article from the literature and present a critique of this article to the group and faculty (Journal Club). Fellows were guided in the preparation of this presentation by one of the critical care medicine faculty. Finally, a written analysis and critique of the article was performed by each fellow. MEASUREMENTS AND MAIN RESULTS: A test was given to each fellow at the beginning and end of the academic year. This test consisted of two pairs of articles on therapy for acute lung injury. For the pretest, each fellow was assigned, at random, one pair of articles. Fellows were given 1 hr to review both articles and to fill out a six-point test to assess their ability and confidence to appraise each article. At the end of the year, each fellow was tested on the opposite pair, the tests were graded in a blinded fashion and the results of each test were compared. Six fellows completed both pre- and posttests. These paired results were analyzed separately, whereas results for another six fellows were conducted as an unpaired analysis. Mean scores increased both for the paired analysis (4.1+/-0.7 vs. 5.1+/-0.5; p = .015) and for the unpaired analysis (4.3+/-0.6 vs. 5.0+/-0.5; p = .012). Self-reported confidence in critical appraisal also increased (2.5+/-0.5 vs. 3.9+/-0.7; p = .004 and 2.6+/-0.5 vs. 3.9+/-0.6; p < .001, respectively). CONCLUSION: Critical appraisal exercises used in the training of critical care medicine fellows appear to improve both ability and confidence to appraise relevant medical literature.

Critical Care↗

Phosphorylation of brush border myosin at threonine on its 20 kDa light chains by a calmodulin-independent kinase activates its ATPase.

A calmodulin-independent kinase isolated from chicken intestinal brush border phosphorylates brush border myosin mainly at an apparently single threonine on its 20 kDa light chains. Phosphorylation to 1.9 mol phosphate/mol myosin activated the myosin actin-activated ATPase about 12-fold, to about 100 nmol/min per mg. Brush border myosin ATPase can thus be activated by phosphorylation either at threonine, by calmodulin-independent kinase, or at serine, by calmodulin-dependent myosin light chain kinase, as previously shown [(1987) FEBS Lett. 223, 262-266].

Actins↗

Purification and characterization of a calmodulin-dependent myosin heavy chain kinase from intestinal brush border.

A calcium- and calmodulin-dependent kinase that represents the majority of the myosin heavy chain kinase activity in chicken intestinal brush borders has been highly purified. The purification steps include gel filtration, high performance chromatography on anion and cation exchangers, and affinity chromatography on calmodulin-Sepharose. The purified kinase consists of a single major, apparently autophosphorylatable polypeptide of 50,000 daltons. The Stokes radius (68 A) and sedimentation coefficient (17.5 S) indicate that it has a molecular weight of approximately 490,000. The kinase catalyzed the incorporation of a maximum of 0.8 mol of phosphate/mol of heavy chain, and essentially no phosphate was incorporated into the light chains. This kinase is distinct from other myosin kinases, but has a number of properties in common with the type II calmodulin-dependent protein kinases.

Animals↗

Phosphorylation of brush border myosin by brush border calmodulin-dependent myosin heavy and light chain kinases.

Calmodulin-dependent myosin light chain kinase isolated from chicken intestinal brush border phosphorylates brush border myosin at an apparently single serine identical to that phosphorylated by smooth muscle myosin light chain kinase. Phosphorylation to 1.8 mol phosphate/mol myosin activated the myosin actin-activated ATPase about 10-fold, to about 50 nmol/min per mg. Myosin phosphorylated on its light chains could then be further phosphorylated to a total of 3.2 mol phosphate per mol by brush border calmodulin-dependent heavy chain kinase. Heavy chain phosphorylation did not alter the actin-activated ATPase of either myosin prephosphorylated on its light chains or of unphosphorylated myosin.

Actins↗

Brush border myosin heavy chain phosphorylation is regulated by calcium and calmodulin.

Myosin from chicken intestinal brush borders is phosphorylated on its heavy chains at threonine by a kinase isolated from brush borders. In contrast to other heavy chain kinases, the brush border kinase activity is dependent on calcium and calmodulin. The partially purified preparation also phosphorylated myosin on its light chains at serine, but in a calmodulin-independent manner. Phosphorylation of the light chains in the absence of calmodulin or both heavy and light chains in the presence of calmodulin activated its actin-activated ATPase activity about 10-fold, to about 50 nmol/min per mg.

Adenosine Triphosphatases↗

Quantitation of proteins on Coomassie blue-stained polyacrylamide gels based on spectrophotometric determination of electroeluted dye.

A procedure for quantitating proteins on Coomassie blue-stained polyacrylamide gels is presented. The method is based on the observations that the dye is rapidly eluted electrophoretically from stained protein bands or spots in the presence of sodium dodecyl sulfate, and that the eluted dye is nondialyzable. Protein may therefore be assayed indirectly by measuring the dye in electroeluents spectrophotometrically. Moreover, the stain elutes more rapidly than the protein, allowing separate recovery of the protein for further analysis. The assay is independent of band or spot size, and does not involve physical disruption of the gel piece or any chemical treatment harsher than the staining process itself. The technique has been applied to the contractile proteins myosin, actin, and commercially obtained standards resolved by one-dimensional electrophoresis, and to proteins in nuclear extracts of HeLa cells on two-dimensional gels.

Animals↗