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Biomedical subjects

L R Jones

Publications and source records attributed to L R Jones.

At least 145 records · Page 8Linked to original sources

Inside the hidden mental health network. Examining mental health care delivery of primary care physicians.

Mental disorder diagnoses among 51 patients, made by a group of 20 family physicians, were compared with diagnoses generated by the Diagnostic Interview Schedule (DIS). Processes of diagnosis, decision making, and treatment planning were then examined through structured physician interviews and chart audits. In this study, 75 of 94 DIS diagnoses (79%) were undetected. During interview and chart audit, the physicians were found to have consistently underestimated, misinterpreted, or neglected psychiatric aspects of care among a majority of patients in the study. These physicians had all satisfactorily completed a psychiatry curriculum designed for family physicians. Analysis of these results suggests that a mental health role is often not integrated into primary care practice, regardless of physician performance during psychiatric training experiences. Assumption of this role appears to be state dependent on involvement with a psychiatric treatment setting. Primary care practice patterns do not seem to result in application of appropriate skills and therapeutic attitudes to detect, diagnose, and correctly manage the majority of mental disorders that occur. The need is reaffirmed for active collaboration between mental health professional and primary care providers in training and in incorporation of psychiatric skills into primary care practice.

Adult↗

Immunoelectron microscopical localization of phospholamban in adult canine ventricular muscle.

The subcellular distribution of phospholamban in adult canine ventricular myocardial cells was determined by the indirect immunogold-labeling technique. The results presented suggest that phospholamban, like the Ca2+-ATPase, is uniformly distributed in the network sarcoplasmic reticulum but absent from the junctional portion of the junctional sarcoplasmic reticulum. Unlike the Ca2+-ATPase, but like cardiac calsequestrin, phospholamban also appears to be present in the corbular sarcoplasmic reticulum. Comparison of the relative distribution of phospholamban immunolabeling in the sarcoplasmic reticulum with that of the sarcolemma showed that the density of phospholamban in the network sarcoplasmic reticulum was approximately 35-fold higher than that of the cytoplasmic side of the sarcolemma, which in turn was found to be three- to fourfold higher than the density of the background labeling. However, a majority of the specific phospholamban labeling within 30 nm of the cytoplasmic side of the sarcolemma was clustered and present over the sarcoplasmic reticulum in the subsarcolemmal region of the myocardial cells, suggesting that phospholamban is confined to the junctional regions between the sarcolemma and the sarcoplasmic reticulum, but absent from the nonjunctional portion of the sarcolemma. Although the resolution of the immunogold-labeling technique used (60 nm) does not permit one to determine whether the specific labeling within 30 nm of the cytoplasmic side of the sarcolemma is associated with the sarcolemma and/or the junctional sarcoplasmic reticulum, it is likely that the low amount of labeling in this region represents phospholamban associated with sarcoplasmic reticulum. These results suggest that phospholamban is absent from the sarcolemma and confined to the sarcoplasmic reticulum in cardiac muscle.

Animals↗

Characteristics of frozen colostrum thawed in a microwave oven.

Use of a microwave oven to thaw frozen colostrum was evaluated. Colostrum was collected from nine cows, four of which were immunized to produce specific colostral antibodies. Colostrum from each cow was frozen, subsequently thawed, and pooled. One-liter aliquots of the pooled colostrum were frozen and assigned randomly to three thawing treatments. Colostrum was thawed using one of three regimens: 10 min in a microwave oven at full power (650 W), 17 min in a microwave oven at half power (325 W), and 25 min in 45 degrees C water. Colostrum thawed in the microwave oven was slightly coagulated and had lower volume and total protein content than colostrum thawed in water. Casein and pH were not different among treatments. Both concentration and total content of immunoglobulin A were higher in the control than in microwave treatments. Neither amount nor concentration of immunoglobulin G and immunoglobulin M were different among treatments. Immunological activity, measured by a hemolytic test, was lower for microwave treatments than the control but did not differ between microwave treatments. Frozen colostrum thawed in a microwave oven should provide a reasonable source of colostrum when fresh high quality colostrum is not available.

Animals↗

Educating family physicians to care for the chronically mentally ill.

More than 50 percent of the chronically mentally ill receive their medical, psychiatric, and social support services from primary care physicians in the general health sector. Despite this high level of involvement with these patients, the majority of family physicians consider their training in the management of patients with mental disorders to be inadequate. This paper describes six categories of critical competencies that should be included in the mental health curricula of family physician training programs: therapeutic attitudes and skills, diagnosis and differential diagnosis, functional assessment, psychopharmacology, management of emergencies, and psychosocial treatments. It outlines the manner in which specific competencies could be incorporated in medical school, in family practice residency training, and in postgraduate continuing medical education as well as the specific elements included in each. The discussion is based on the assumption that more effective participation by family physicians in the treatment of chronic psychiatric illness requires active attention throughout the continuum of medical education.

Adult↗

Developmental changes in cardiac sarcoplasmic reticulum in sheep.

Physiologic studies suggest that the myocardium from fetal and newborn sheep functions at a higher contractile state with decreased contractile reserve when compared to the myocardium of adult sheep. To investigate the role of Ca2+ transport by the sarcoplasmic reticulum (SR) in this phenomenon, we studied functional properties and protein composition of cardiac SR vesicles isolated from fetal and maternal sheep. Active accumulation of Ca2+ and the density of the Ca2+ pump protein were decreased 60% (p less than 0.01) in fetal SR vesicles; however Ca2+-dependent ATPase activity was decreased only 30% (p less than 0.01). This decreased difference in Ca2+-dependent ATPase activities was accounted for by the higher turnover number measured for the Ca2+ pump of fetal SR vesicles (1.6-fold increased, p less than 0.01). Ryanodine, an alkaloid which blocks Ca2+ efflux from cardiac SR vesicles, stimulated Ca2+ uptake more effectively in fetal SR vesicles, suggesting that these vesicles had a higher passive Ca2+ permeability during conditions of active Ca2+ transport. Protein compositional studies showed that the content of phospholamban was decreased in fetal SR vesicles and was correlated with the decrease in the density of Ca2+ pumps. In contrast, the content of calsequestrin and the density of [3H]nitrendipine-binding sites were increased approximately 2-fold in fetal SR vesicles. These functional and compositional differences between SR vesicles isolated from fetal and maternal sheep may indicate that there is relatively more junctional SR in fetal hearts. Since the SR regulates muscle contraction by modulating intracellular Ca2+ concentration, it is possible that developmental alterations in cardiac SR may contribute to the decreased myocardial contractile reserve noted in fetal sheep.

Animals↗

Sequence analysis of phospholamban. Identification of phosphorylation sites and two major structural domains.

Phospholamban is a regulatory protein in cardiac sarcoplasmic reticulum that is phosphorylated by cAMP- and Ca2+/calmodulin-dependent protein kinase activities. In this report, we present the partial amino acid sequence of canine cardiac phospholamban and the identification of the sites phosphorylated by these two protein kinases. Gas-phase protein sequencing was used to identify 20 NH2-terminal residues. Overlap peptides produced by trypsin or papain digestion extended the sequence 16 residues to give the following primary structure: Ser-Ala-Ile-Arg-Arg-Ala-Ser-Thr-Ile-Glu-Met-Pro-Gln-Gln-Ala- Arg-Gln-Asn-Leu-Gln-Asn-Leu-Phe-Ile-Asn-Phe-(Cys)-Leu-Ile-Leu-Ile-(Cys)- Leu-Leu-Leu-Ile-. Phospholamban phosphorylated by either cAMP-dependent or Ca2+/calmodulin-dependent protein kinase was cleaved with trypsin, and the major phosphorylated peptide (comprising greater than 70% of the incorporated 32P label) was purified by reverse-phase high performance liquid chromatography. The identical sequence was revealed for the radioactive peptide obtained from phospholamban phosphorylated by either kinase: Arg-Ala-Ser-Thr-Ile-Glu-Met-Pro-Gln-Gln-. The adjacent residues Ser7 and Thr8 of phospholamban were identified as the unique sites phosphorylated by cAMP- and Ca2+/calmodulin-dependent protein kinases, respectively. These results establish that phospholamban is an oligomer of small, identical polypeptide chains. A hydrophilic, cytoplasmically oriented NH2-terminal domain on each monomer contains the unique, adjacent residues phosphorylated by cAMP- and Ca2+/calmodulin-dependent protein kinase activities. Analysis by hydropathic profiling and secondary structure prediction suggests that phospholamban monomers also contain a hydrophobic domain, which could form amphipathic helices sufficiently long to traverse the sarcoplasmic reticulum membrane. A model of phospholamban as a pentamer is presented in which the amphipathic alpha-helix of each monomer is a subunit of the pentameric membrane-anchored domain, which is comprised of an exterior hydrophobic surface and an interior hydrophilic region containing polar side chains.

Amino Acid Sequence↗

Evidence for the presence of phospholamban in the endoplasmic reticulum of smooth muscle.

In microsomal vesicles isolated from several smooth muscles many polypeptides were phosphorylated by the catalytic subunit of cyclic AMP-dependent protein kinase. In pig stomach and in rabbit and dog aorta components of Mr 22,000 and 11,000 were identified as forms of phospholamban. These polypeptides were, however, not observed in pig aorta. These phospholamban-like polypeptides presented the same electrophoretic mobility in sodium dodecyl sulphate gels as cardiac phospholamban, and the 22,000 Mr form showed a similar reaction to heat treatment in sodium dodecyl sulphate. Antibodies against purified canine cardiac phospholamban cross-reacted with the 22,000 and 11,000 Mr phosphorylatable polypeptides from smooth muscle membranes. Subcellular fractionation of porcine stomach smooth muscle indicated that phospholamban was present in the membranes of the endoplasmic reticulum and not in the plasma membranes. Phospholamban was also phosphorylated by an endogenous calcium-calmodulin-dependent protein kinase and by an endogenous cyclic AMP-dependent kinase. It is concluded that the endoplasmic reticulum of many, but possibly not all, smooth muscles contains phospholamban. However, the physiological role of phospholamban in smooth muscle remains to be established.

Animals↗

Proteolytic cleavage of phospholamban purified from canine cardiac sarcoplasmic reticulum vesicles. Generation of a low resolution model of phospholamban structure.

Purified phospholamban isolated from canine cardiac sarcoplasmic reticulum vesicles was subjected to proteolysis and peptide mapping to localize the different sites of phosphorylation on the protein and to gain further information on its subunit structure. Five different proteases (trypsin, papain, chymotrypsin, elastase, and Pronase) degraded the oligomeric 27-kDa phosphoprotein into a major 21-22-kDa protease-resistant fragment. No 32P was retained by this protease-resistant fragment, regardless of whether phospholamban had been phosphorylated by cAMP-dependent protein kinase, Ca2+/calmodulin-dependent protein kinase, or protein kinase C. Phosphoamino acid analysis and thin-layer electrophoresis of liberated phosphopeptides revealed that 1 threonine and 2 serine residues were phosphorylated in phospholamban and that 1 of these serine residues and the threonine residue were in close proximity. Only serine was phosphorylated by cAMP-dependent protein kinase, whereas Ca2+-calmodulin-dependent protein kinase phosphorylated exclusively threonine. The results demonstrate that phospholamban has a large protease-resistant domain and a smaller protease-sensitive domain, the latter of which contains all of the sites of phosphorylation. The 21-22-kDa protease-resistant domain, although devoid of incorporated 32P, was completely dissociated into identical lower molecular weight subunits by boiling in sodium dodecyl sulfate, suggesting that this region of the molecule promotes the relatively strong interactions that hold the subunits together. The data presented lend further support for a model of phospholamban structure in which several identical low molecular weight subunits are noncovalently bound to one another, each containing one site of phosphorylation for cAMP-dependent protein kinase and another site of phosphorylation for Ca2+/calmodulin-dependent protein kinase.

Animals↗

Localization of phospholamban in slow but not fast canine skeletal muscle fibers. An immunocytochemical and biochemical study.

Phospholamban, originally described as a cardiac sarcoplasmic reticulum protein, was localized in cryostat sections of three adult canine skeletal muscles (gracilis, extensor carpi radialis, and superficial digitalis flexor) by immunofluorescence labeling with highly specific phospholamban antibodies. Only some myofibers were strongly labeled with phospholamban antibodies. The labeling of myofibers with phospholamban antibodies was compared to the distribution of Type I (slow) and Type II (fast) myofibers as determined by staining adjacent sections cytochemically for the alkali-stable myosin ATPase, a specific marker for Type II myofibers. All the skeletal myofibers labeled for phospholamban above background levels corresponded to Type I (slow) myofibers. The presence of phospholamban in microsomal fractions isolated from canine superficial digitalis flexor (89 +/- 3% Type I) and extensor carpi radialis skeletal muscle (14 +/- 6% Type I) was confirmed by immunoblotting. Antiserum to cardiac phospholamban bound to proteins of apparent Mr values of 25,000 (oligomeric phospholamban) and 5,000-6,000 (monomeric phospholamban) in sarcoplasmic reticulum vesicles from both muscles. Quantification of phospholamban in sarcoplasmic reticulum vesicles from cardic, slow, and fast skeletal muscle tissues following phosphorylation with [gamma-32P] ATP suggested that superficial digitalis flexor and extensor carpi radialis skeletal muscle contained about 16 and 3%, respectively, as much phospholamban as cardiac muscle per unit of sarcoplasmic reticulum. The presence of phospholamban in both Type I (slow) and cardiac muscle fibers supports the possibility that the Ca2+ fluxes across the sarcoplasmic reticulum in both fiber types are similarly regulated, and is consistent with the idea that the relaxant effect of catecholamines on slow skeletal muscle is mediated in part by phosphorylation of phospholamban.

Animals↗

Influence of age, sex, lactational state and exogenous growth hormone on erythrocyte UMP synthase in dairy cattle.

UMP synthase activity was determined in erythrocytes of Holstein cattle to assess differences attributable to age, sex, lactational state, and exogenous growth hormone. Newborns had 80% more UMP synthase activity per ml erythrocytes than mature cattle. Males had 10% more UMP synthase activity than females of the same age. Lactating and non-lactating mature females did not differ in UMP synthase activity. Over the first six weeks of life, the decrease in UMP synthase activity was less pronounced in calves partially deficient for the enzyme. Ten daily injections of growth hormone altered neither UMP synthase activity in erythrocytes nor orotic acid concentrations in milk and urine.

Aging↗

Controllership in the public sector.

Ineffective or incompetent controllership is, perhaps, the most common managerial failure found in the public sector. This failure affects outcomes and achievements in every area of public policy--often profoundly. Controllers design and operate management control systems. The effectiveness of alternative control system designs depends upon the cost and production behavior of the good or service in question. But control strategies are seldom consciously or effectively matched to circumstances. As a consequence, public programs are too often simultaneously overcontrolled and out of control.

Budgets↗

Identification of an endogenous protein kinase C activity and its intrinsic 15-kilodalton substrate in purified canine cardiac sarcolemmal vesicles.

The cardiac sarcolemmal 15-kDa protein, previously shown to be the principal sarcolemmal substrate phosphorylated in intact heart in response to beta-adrenergic stimulation (Presti, C. F., Jones, L. R., and Lindemann J. P. (1985) J. Biol. Chem. 260, 3860-3867), was demonstrated to be the major substrate phosphorylated in purified canine cardiac sarcolemmal vesicles by an intrinsic protein kinase C activity. The intrinsic protein kinase C, detected by its ability to phosphorylate H1 histones, was most concentrated in cardiac sarcolemmal vesicles and absent from sarcoplasmic reticulum membranes. Unmasking techniques localized the intrinsic protein kinase activity and its principal endogenous substrate, the 15-kDa protein, to the cytoplasmic surfaces of sarcolemmal vesicles; phospholamban contaminating the sarcolemmal preparation was not significantly phosphorylated. The intrinsic protein kinase C required micromolar Ca2+ for activity, but not calmodulin. Half-maximal phosphorylation of the 15-kDa protein occurred at 10 microM Ca2+; optimal phosphorylation of the 15-kDa protein by protein kinase C and Ca2+ was additive to that produced by cAMP-dependent protein kinase. Exogenous phospholipids were not required to activate endogenous protein kinase C. However, heat-treated sarcolemmal vesicles, in which intrinsic protein kinase activities were inactivated, were sufficient to maximally activate soluble protein kinase C prepared from rat brain, suggesting that all the necessary phospholipid cofactors were already present in sarcolemmal vesicles. Of the many proteins present in sarcolemmal vesicles, only the 15-kDa protein was phosphorylated significantly in heat-inactivated sarcolemmal vesicles by soluble protein kinase C, confirming that the 15-kDa protein was a preferential substrate for this enzyme. Consistent with a protein kinase C activity in sarcolemmal vesicles, the tumor-promoting phorbol ester 12-O-tetradecanoylphorbol 13-acetate stimulated 15-kDa protein phosphorylation severalfold, producing approximately 70% of the maximal phosphorylation even in the absence of significant ionized Ca2+. The results are compatible with an intrinsic protein kinase C activity in sarcolemmal vesicles whose major substrate is the 15-kDa protein.

Animals↗

Purification and characterization of phospholamban from canine cardiac sarcoplasmic reticulum.

Phospholamban, the putative protein regulator of the Ca2+ pump of cardiac sarcoplasmic reticulum, was purified to apparent homogeneity from canine cardiac sarcoplasmic reticulum vesicles by selective extraction with sodium cholate, followed by adsorption to calcium oxalate, solubilization in Zwittergent 3-14, and specific elution from p-hydroxymercuribenzoate-agarose. Phospholamban, isolated in the dephosphorylated state, was purified 80-fold in 15% yield (approximately 2 mg of phospholamban/g of sarcoplasmic reticulum protein). Nondissociated phospholamban exhibited an apparent Mr = 25,000 in sodium dodecyl sulfate-polyacrylamide gels. Partially dissociated phospholamban, induced by boiling in sodium dodecyl sulfate, exhibited five distinct mobility forms in sodium dodecyl sulfate-polyacrylamide gels, of apparent molecular weights between 5,000-6,000 and 25,000. Phospholamban was phosphorylated to a level of 190 nmol of Pi/mg of protein by cAMP-dependent protein kinase, consistent by minimum stoichiometry with a subunit molecular weight of approximately 5,000. Phospholamban prepared by the present method was different in several respects from the proteins that have been isolated in other laboratories. Pure phospholamban was cysteine rich, containing 6 residues/100 amino acid residues. Dephosphorylated phospholamban was strongly basic with a pI = 10; phosphorylation decreased the pI to approximately 6.7. Pure phospholamban (and phospholamban present in sarcoplasmic reticulum vesicles) was not readily extracted into acidified chloroform/methanol, suggesting that the protein does not behave as an acidic proteolipid. The purified protein was highly antigenic. Phospholamban was localized by immunochemical methods to cardiac membranes enriched in sarcoplasmic reticulum, but was absent from sarcoplasmic reticulum membranes prepared from fast skeletal muscle. The method described for isolation of cardiac phospholamban is highly reproducible and relatively simple, and should be useful for further detailed studies designed to probe the molecular structure of the protein.

Adenosine Triphosphatases↗

Demonstration of a Na+/H+ exchange activity in purified canine cardiac sarcolemmal vesicles.

Purified canine cardiac sarcolemmal membrane vesicles exhibit a sodium ion for proton exchange activity (Na+/H+ exchange). Na+/H+ exchange was demonstrated both by measuring rapid 22Na uptake into sarcolemmal vesicles in response to a transmembrane H+ gradient and by following H+ transport in response to a transmembrane Na+ gradient with use of the probe acridine orange. Maximal 22Na uptake into the sarcolemmal vesicles (with starting intravesicular pH = 6 and extravesicular pH = 8) was approximately 20 nmol/mg protein. The extravesicular Km of the Na+/H+ exchange activity for Na+ was determined to be between 2 and 4 mM (intravesicular pH = 5.9, extravesicular pH = 7.9), as assessed by measuring the concentration dependence of the 22Na uptake rate and the ability of extravesicular Na+ to collapse an imposed H+ gradient. All results suggested that Na+/H+ exchange was reversible and tightly coupled. The Na+/H+ exchange activity was assayed in membrane subfractions and found most concentrated in highly purified cardiac sarcolemmal vesicles and was absent from free and junctional sarcoplasmic reticulum vesicles. 22Na uptake into sarcolemmal vesicles mediated by Na+/H+ exchange was dependent on extravesicular pH, having an optimum around pH 9 (initial internal pH = 6). Although the Na+/H+ exchange activity was not inhibited by tetrodotoxin or digitoxin, it was inhibited by quinidine, quinacrine, amiloride, and several amiloride derivatives. The relative potencies of the various inhibitors tested were found to be: quinacrine greater than quinidine = ethylisopropylamiloride greater than methylisopropylamiloride greater than dimethylamiloride greater than amiloride. The Na+/H+ exchange activity identified in purified cardiac sarcolemmal vesicles appears to be qualitatively similar to Na+/H+ exchange activities recently described in intact cell systems. Isolated cardiac sarcolemmal vesicles should prove a useful model system for the study of Na+/H+ exchange regulation in myocardial tissue.

Alamethicin↗

Isoproterenol-induced phosphorylation of a 15-kilodalton sarcolemmal protein in intact myocardium.

The effect of beta-adrenergic stimulation on sarcolemmal protein phosphorylation was examined in intact ventricular myocardium. Isolated guinea pig ventricles were perfused via the coronary arteries with 32Pi after which membrane vesicles enriched 3-5-fold in sarcolemma were isolated by differential centrifugation followed by sucrose gradient centrifugation. Perfusion of hearts with isoproterenol stimulated 32P incorporation into a protein of apparent molecular weight of 15,000, which copurified with sarcolemmal vesicles. The increase in 32P incorporation was rapid in onset and elevated 2.5-3.0-fold after 30-45 s exposure of hearts to 100 nM isoproterenol. A positive correlation was found between stimulation of phosphorylation of the 15-kDa protein and the increase in the maximal rate of developed tension in intact ventricles after administration of isoproterenol. Phosphorylated phospholamban (most likely present as a contaminant) was also identified in the same sarcolemmal preparations. However, phospholamban and the 15-kDa sarcolemmal substrate were different proteins. Boiling of the membrane samples in sodium dodecyl sulfate prior to electrophoresis dissociated the high Mr form of phospholamban into the form of lower Mr but did not alter the mobility of the 15-kDa protein in sodium dodecyl sulfate-polyacrylamide gels. The 15-kDa protein did not undergo the electrophoretic mobility shift that is characteristic of phospholamban after cAMP-dependent phosphorylation nor did it cross-react with a highly specific phospholamban antibody. In vitro phosphorylation experiments conducted with the unmasking agent Triton X-100 suggested that the 15-kDa protein was localized to the cytoplasmic surfaces of sarcolemmal vesicles. These results demonstrate phosphorylation of a sarcolemmal protein, distinct from phospholamban, in response to beta-adrenergic stimulation of the heart. Phosphorylation of the sarcolemmal 15-kDa protein may play a role in mediating the effects of beta-adrenergic agonists on cardiac contractile force.

Animals↗

Assessment in humans of hypolipidemia induced by orotic acid.

The hypolipidemia induced by oral orotic acid in rats but not in other experimental species was investigated in human adults. Twelve outpatients in a medically-supervised weight reduction program participated in the study. After adaptation to their restricted dietary regime for at least 8 weeks, all subjects were given placebos (1 g lactose/day) for 2 weeks, then orotic acid (1 g/day) for 4 weeks, followed by placebos for 2 more weeks. Pertinent dietary, urinary, and serum parameters were monitored biweekly and intake of calories, carbohydrate, protein and fat was similar throughout. The ingestion of orotic acid did not lower serum triglycerides or cholesterol in these subjects. Since the daily dose tested corresponds to the orotic acid in 12 liters of cow's milk, milk orotic acid at levels normally consumed should not be considered a hypolipidemic or hypocholesteremic agent for humans.

Adult↗

Erythrocyte ribosyluric acid of dairy cattle normal and deficient for uridine monophosphate synthase.

An inherited deficiency of uridine-5'-monophosphate synthase has recently been discovered in dairy cattle. The possibility was explored that this deficiency might be detected by low amounts of erythrocyte 3-N-ribosyluric acid, since it is synthesized by uridine monophosphate synthase. Ribosyluric acid was lower in six dairy cows with the deficiency compared to six controls, paired on the basis of age, stage of lactation, and diet (.66 vs. 1.38 mumol/ml). However, ribosyluric acid concentration measured in 50 normal cows showed a wide range (.28 to 2.35 mumol/ml), with nine lower than the mean for the deficient animals. This extensive variability may limit the usefulness of ribosyluric acid in diagnosing the inherited deficiency of uridine monophosphate synthase.

Animals↗