Thesis (M.S.)--University of Missouri-Columbia, 2004. / Typescript. Vita. Includes bibliographical references (leaf 58). Also available on the Internet.

Thesis (M.S.)--University of Michigan, 1994. / eContent provider-neutral record in process. Description based on print version record.

Thesis (M.S.)--University of Missouri-Columbia, 2004. / Typescript. Vita. Includes bibliographical references (leaf 58). Also available on the Internet.

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Calcium transport across the placenta is asymmetrical and is believed to be an active transport. An essential step in such a transport is translocation of the ion across a single plasma membrane. The objective of this thesis was to study the Ca2+ -related properties of the placental plasma membranes and to gain some knowledge of their role in Ca2+ -transport. Three Ca2+ -related properties were studied: 1. Ca2+ -binding to the placental plasma membranes; 2. The membrane bound enzyme Ca -ATPase; and 3. Ca2+ -uptake by the placental plasma membrane vesicles. Ca2+ -binding properties of the membrane preparation were studied by the use of a new method, the flow dialysis system. Two types of sites for Ca were found: 1) high affinity, low capacity sites, and 2) low affinity, high capacity sites. The high affinity sites had 10-fold higher affinity for Ca2+ than for Mg2+ . A calcium-stimulated, membrane-bound enzyme, namely Ca2+ -ATPase, was located in the placental plasma membranes. This enzyme is distinct from the Na+, K+-ATPase and alkaline phosphatase. The enzyme can be activated by Mg2+ but with lower efficiency. Both Ca2+ and Mg2+ activate the enzyme at the same site. A formula was derived, enabling one to predict very precisely the velocity of the enzyme incubated under any combination of Ca2+ and Mg2+ ; this relationship is presented in a three dimensional model. The formula can be used for other enzymes or other substrates, as was demonstrated with ATP and ADP. The placental plasma membrane vesicles are capable of accumulating Ca2+ . Ca2+ -uptake was defined as the amount of Ca2+ which is not available for rapid exchange and cannot be displaced by a high concentration of competitor in the presence of ATP. This definition is different from and more accurate than the one which is widely used and cited in the literature. An intravesicular Ca2+ concentration of 190 mM was recorded, which was 24-fold higher than the external Ca2+ concentration (8 mM). Ca2+ -uptake was dependent on ATP hydrolysis by the placental Ca2+ -ATPase. This process was independent of Mg2+ . It is suggested that while the substrate for Ca2+ -ATPase is Ca-ATP, the substrate for Ca2+ -uptake is Ca2+. The overall Ca2+ -related properties of the placental plasma membranes are independent of Mg and the entire process from binding to membrane through activation of the enzyme and finally Ca2+ -uptake is dependent on Ca2+ alone. This situation is unique to the placental plasma membranes. It is tempting to speculate that the link between the maternal and the fetal circulation is achieved by forming vesicles loaded with Ca2+ on the maternal side and unloading them through fusion with the basal plasma membrane on the fetal side. The Ca2+ -related properties of placental plasma membranes described in this thesis, provide many answers regarding the first step in the asymmetrical transplacental Ca2+ -transport. Further investigation is required before a full understanding of the entire process is achieved. / Medicine, Faculty of / Cellular and Physiological Sciences, Department of / Graduate

The mechanism of regulation of the Ca²⁺ pump ATPase of the human erythrocyte membrane by calmodulin, cyclic AMP and the anion channel was studied using membrane fragments, resealed "ghosts", inside-out vesicles and a Triton X-100 solubilized enzyme preparation. The (Ca²⁺ + Mg²⁺ )-ATPase activity in erythrocyte membranes or a Triton X-100 solubilized enzyme preparation showed biphasic (high and low affinity) Ca²⁺ activation kinetics. The anionic calcium binding protein, calmodulin, increased both the calcium sensitivity (Kca²⁺) and the maximum velocity (Vmax ) of the enzyme. Certain polyanionic agents (poly-L-aspartic acid, poly-L-glutamic acid), alicyclic sulfonic acids (HEPES,N-2-hydroxyethylpiperazine-N¹-2-ethanesulfonic acid, MES,2-N- (morpholinoethanesulfonic acid)), and aromatic carboxylic acids (benzoic and salicylic acids) increased the Kca²⁺ but not the Vmax of (Ca²⁺ + Mg²⁺ )-ATPase in erythrocyte membranes and Triton X-100 solubilized enzyme preparations. Trifluoperazine (30 μM) antagonized activation of the enzyme by calmodulin and poly-L-aspartic acid, but not by sodium-HEPES or sodium-MES. Limited trypsin proteolysis of (Ca²⁺ + Mg²⁺ )-ATPase in the erythrocyte membrane abolished activation by calmodulin, poly-L-aspartic acid and sodium-HEPES. These results suggest that the modulation of the Ca²⁺ sensitivity of (Ca²⁺ + Mg²⁺ )-ATPase by calmodulin may be associated with the anionic properties of this protein, and that this property can be mimicked by some other anions, probably by interacting at an anion-regulatory site on the enzyme. Cyclic AMP (5 μM) was found to inhibit the (Ca²⁺ + Mg²⁺)-ATPase activity (approx. 20%) in erythrocyte membranes, probably via endogenous cyclic AMP protein kinase, since this effect could be blocked by cyclic AMP protein kinase inhibitor (PKI) from the rabbit skeletal muscle, By contrast, bovine heart PKI stimulated (Ca²⁺ + Mg²⁺ )-ATPase activity (approx. 100%) by increasing the Kca²⁺ but not the Vmax of the enzyme in membrane or Triton X-100 solubilized preparations. At a low calcium concentration the stimulation by bovine heart PKI and saturating levels of calmodulin was additive, suggesting that the two effectors acted by distinct mechanisms. The stimulation of (Ca²⁺ + Mg²⁺ )-ATPase activity by bovine heart PKI was not solely due to its antagonism of the protein kinase because a) modification of arginine residues of bovine heart PKI abolished its inhibition of cyclic AMP protein kinase, but had no effect on the stimulation of (Ca²⁺ + Mg²⁺ )-ATPase; b) trifluoperazine (20 μM) antagonized the stimulation of (Ca²⁺ + Mg²⁺ )-ATPase by PKI, similarly to its antagonism of calmodulin stimulation, but it did not affect the inhibition of protein kinase by PKI. It is suggested that different mechanisms are involved in the inhibition of cyclic AMP protein kinase and the stimulation of (Ca²⁺ + Mg²⁺ )-ATPase by bovine heart cyclic AMP PKI. Next, the role of anion channel blockers on the (Ca²⁺ + Mg²⁺ )- ATPase was studied. The photolabeling reagent N-(4-azido-2-nitrophenyl)- 2 aminoethylsulfonate (NAP-taurine) was found to inhibit the (Ca²⁺+ Mg²⁺ )-ATPase of fragmented red cell membranes. Half maximal inhibition occurred between 25 μM and 50 μM. At these concentrations Mg²⁺ -ATPase and (Na⁺ + K⁺)-ATPase activities in the membranes were not affected. The reversible inhibition of (Ca²⁺ + Mg²⁺ )-ATPase produced by NAP-taurine in the dark became irreversible after photolysis in the presence of this reagent. Incubation of the membranes with Ca²⁺ , Mg²⁺ , ATP or calmodulin, prior to photolysis in the presence of NAP-taurine, did not protect the enzyme from Inhibition. Limited trypsin proteolysis of (Ca²⁺ + Mg²⁺ )-ATPase in fragmented membranes, which abolished activation by calmodulin, did not affect the inhibition by NAP-taurine. NAP-taurine was found to Inhibit the (Ca²⁺ + Mg²⁺ )-ATPase activity from the cytoplasmic side of the membrane, as determined from the following experiments. Addition of NAP-taurine (50 μM) to resealed erythrocyte ghosts inhibited less than 5% of the (Ca²⁺ + Mg²⁺ )-ATPase activity, compared to 50-60% Inhibition in ghosts resealed in the presence of 50 μM NAP-taurine. Furthermore, NAP-taurine inhibited ATP-dependent Ca²⁺ - transport into inside-out vesicles at a similar concentration (50 μM). The inhibition of the (Ca²⁺ + Mg²⁺ )-ATPase activity of membranes by NAP-taurine appeared to be a direct action on the enzyme, rather than through inhibition of the anion channel, as (Ca²⁺ + Mg²⁺ )-ATPase activity was not inhibited in membranes made from red blood cells reacted irreversibly with 50 μM NAP-taurine or the anion channel blocker 4,4'-diisothiocyano- 2,2' stilbene disulfonate (DIDS) (5 μM) or in membranes assayed in the presence of another anion channel blocker, probenecid (125 μM). This is the first reported selective antagonist of the Ca²⁺ pump, and it is suggested that NAP-taurine could be a useful tool for studying the Ca²⁺- transport ATPase in a variety of cells. / Pharmaceutical Sciences, Faculty of / Graduate

Ph. D. / incomplete_metadata

The purpose of this retrospective study was to determine the effectiveness of an innovative unit-wide phosphorus football contest and nutrition education intervention directed at improving the serum phosphorus levels and calcium-phosphorus products of patients undergoing routine hemodialysis. Patients at the Clarian Health Partners dialysis center located at 2140 North Capitol Avenue in Indianapolis, Indiana participated in the "National Fosphorus League Phootball" contest, a theme game that allowed patients to join a team and compete against other teams in the dialysis center over a four month time period from September 2005 to December 2005. Additional nutrition education was also provided to all the hemodialysis patients during the months of the phosphorus football contest. Identical patient information from September 2004 to December 2004 was also collected for baseline comparison of serum phosphorus levels and calcium-phosphorus products when only routine education and instruction was provided.Pearson Chi-Square analyses and a series of three-way ANOVAs were performed on the data collected. Overall, it was determined that patients who participated in the phosphorus football contest and received Vitamin D therapy were potentially two times likely to have serum phosphorus levels and calcium-phosphorus products in the goal ranges. / Department of Family and Consumer Sciences

The purpose of this study was to analyze the effect of fatty acid chain length and quantity on the bioavailability of calcium. Thirteen healthy subjects were randomly assigned to a series of 5 test meals containing varying types and levels of fat and calciumconsumed over a three week time period. The test meals included 10 grams MCT oil (MCT 10), 20 grams MCT oil (MCT 20), 10 grams beef fat (BT 10), 20 grams beef fat (BT 20), and calcium only (Ca). Calcium absorption was assessed using timed urine collections following a specified calcium load. Three day food records were obtained to assess typical nutrient intakes of the subjects coming into and during the study. MCT oil provided better absorption of the calcium supplement than did the beef tallow. A difference was also noted in the absorption of calcium based on the amount of fat consumed. A higher intake of MCT oil (10 g vs. 20 g) appeared to favor the absorption of calcium. Urine calcium excretion was significantly greater (p < .009) during the MCT oil treatments (MCT 10, MCT 20) compared to the beef fat treatments (BT 10, BT 20), suggesting reduced calcium absorption during the beef fat treatments. There were no differences in mean calcium excretion based on quantity of fat consumed ( 10 g vs. 20 g), nor any interaction between type of fat and amount. Tests for detecting differences between individual treatments indicated a significance difference (p < .05) in calcium excretion between MCT 20 and BT 10 treatments. Urine calcium excretion was corrected for body size using urine calcium/creatinine ratio (Ca/Cr). There was a significant time effect between the 0 - 2, 2 - 4 hour time periods (p < .005) and the various treatments for Ca/Cr. Though not significant, mean Ca/Cr was highest for the calcium treatment (0.42), compared to the MCT oil treatments (36, z of MCT 10 & MCT 20), and beef fat treatments (28, x of BT 10 & BT 20). The beef fat treatments significantly decreased the absorption of calcium compared to the MCT oil treatments. It appears that beef fat, when compared to the calcium only treatment, decreased calcium absorption. / Department of Home Economics