Characterising cement-superplasticiser interaction using zeta potential measurements

The zeta potential generated at the interface between cement particle surfaces adsorbed with superplasticisers have been studied using electroacoustic technique, which is capable of measuring zeta potential at high concentrated suspensions. The study has been undertaken to examine the differences in the magnitude of the zeta potential for ordinary Portland cement (OPC) and Portland pozzolanic (fly ash) cement (PPC) pastes along with the differential impacts of different types of superplasticisers on both the varieties of cement pastes. In the latter context, the effects of three different types of superplasticisers namely Ligno Sulphonate (LS), Sulphonated Melamine Formaldehyde (SMF) and Sulphonated Naphthalene Formaldehyde (SNF) have been specifically studied. The results show that the cement pastes with PPC shows better dispersion when compared with the OPC. The paper also endeavors to unfold the relationship and significance of cement interaction with three different superplasticisers.

A calcium-dependent interaction between calmodulin and the calponin homology domain of human IQGAP1

IQGAPs are cytoskeletal scaffolding proteins which collect information from a variety of signalling pathways and pass it on to the microfilaments and microtubules. There is a well-characterised interaction between IQGAP and calmodulin through a series of IQ-motifs towards the middle of the primary sequence. However, it has been shown previously that the calponin homology domain (CHD), located at the N-terminus of the protein, can also interact weakly with calmodulin. Using a recombinant fragment of human IQGAP1 which encompasses the CHD, we have demonstrated that the CHD undergoes a calcium ion-dependent interaction with calmodulin. The CHD can also displace the hydrophobic fluorescent probe 1-anilinonaphthalene-8-sulphonate from calcium-calmodulin, suggesting that the interaction involves non-polar residues on the surface of calmodulin. Molecular modelling identified a possible site on the CHD for calmodulin interaction. The physiological significance of this interaction remains to be discovered.

FhCaBP3: A Fasciola hepatica calcium binding protein with EF-hand and dynein light chain domains

A DNA sequence encoding a protein with predicted EF-hand and dynein light chain binding domains was identified in a Fasciola hepatica EST library. Sequence analysis of the encoded protein revealed that the most similar known protein was the Fasciola gigantica protein FgCaBP3 and so this newly identified protein was named FhCaBP3. Molecular modelling of FhCaBP3 predicted a highly flexible N-terminal region, followed by a domain containing two EF-hand motifs the second of which is likely to be a functioning divalent ion binding site. The C-terminal domain of the protein contains a dynein light chain like region. Interestingly, molecular modelling predicts that calcium ion binding to the N-terminal domain destabilises the ß-sheet structure of the C-terminal domain. FhCaBP3 can be expressed in, and purified from, Escherichia coli. The recombinant protein dimerises and the absence of calcium ions appeared to promote dimerisation. Native gel shift assays demonstrated that the protein bound to calcium and manganese ions, but not to magnesium, barium, zinc, strontium, nickel, copper or cadmium ions. FhCaBP3 interacted with the calmodulin antagonists trifluoperazine, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide and chlorpromazine as well as the myosin regulatory light chain-binding drug praziquantel. Despite sequence and structural similarities to other members of the same protein family from F. hepatica, FhCaBP3 has different biochemical properties to the other well characterised family members, FH22 and FhCaBP4. This suggests that each member of this trematode calcium-binding family has discrete functional roles within the organism.

Comparative capacitative calcium entry mechanisms in canine pulmonary and renal arterial smooth muscle cells.

Experiments were performed to determine whether capacitative Ca(2+) entry (CCE) can be activated in canine pulmonary and renal arterial smooth muscle cells (ASMCs) and whether activation of CCE parallels the different functional structure of the sarcoplasmic reticulum (SR) in these two cell types. The cytosolic [Ca(2+)] was measured by imaging fura-2-loaded individual cells. Increases in the cytosolic [Ca(2+)] due to store depletion in pulmonary ASMCs required simultaneous depletion of both the inositol 1,4,5-trisphosphate (InsP(3))- and ryanodine (RY)-sensitive SR Ca(2+) stores. In contrast, the cytosolic [Ca(2+)] rises in renal ASMCs occurred when the SR stores were depleted through either the InsP(3) or RY pathways. The increase in the cytosolic [Ca(2+)] due to store depletion in both pulmonary and renal ASMCs was present in cells that were voltage clamped and was abolished when cells were perfused with a Ca(2+)-free bathing solution. Rapid quenching of the fura-2 signal by 100 microM Mn(2+) following SR store depletion indicated that extracellular Ca(2+) entry increased in both cell types and also verified that activation of CCE in pulmonary ASMCs required the simultaneous depletion of the InsP(3)- and RY-sensitive SR Ca(2+) stores, while CCE could be activated in renal ASMCs by the depletion of either of the InsP(3)- or RY-sensitive SR stores. Store depletion Ca(2+) entry in both pulmonary and renal ASMCs was strongly inhibited by Ni(2+) (0.1-10 mM), slightly inhibited by Cd(2+) (200-500 microM), but was not significantly affected by the voltage-gated Ca(2+) channel (VGCC) blocker nisoldipine (10 microM). The non-selective cation channel blocker Gd(3+) (100 microM) inhibited a portion of the Ca(2+) entry in 6 of 18 renal but not pulmonary ASMCs. These results provide evidence that SR Ca(2+) store depletion activates CCE in parallel with the organization of intracellular Ca(2+) stores in canine pulmonary and renal ASMCs.

Calcium oscillations in interstitial cells of the rabbit urethra.

Measurements were made (using fast confocal microscopy) of intracellular Ca2+ levels in fluo-4 loaded interstitial cells isolated from the rabbit urethra. These cells exhibited regular Ca2+ oscillations which were associated with spontaneous transient inward currents recorded under voltage clamp. Interference with D-myo-inositol 1,4,5-trisphosphate (IP3) induced Ca2+ release using 100 microm 2-aminoethoxydiphenyl borate, and the phospholipase C (PLC) inhibitors 2-nitro-4-carboxyphenyl N,N-diphenylcarbamate and U73122 decreased the amplitude of spontaneous oscillations but did not abolish them. However, oscillations were abolished when ryanodine receptors were blocked with tetracaine or ryanodine. Oscillations ceased in the absence of external Ca2+, and frequency was directly proportional to the external Ca2+ concentration. Frequency of Ca2+ oscillation was reduced by SKF-96365, but not by nifedipine. Lanthanum and cadmium completely blocked oscillations. These results suggest that Ca2+ oscillations in isolated rabbit urethral interstitial cells are initiated by Ca2+ release from ryanodine-sensitive intracellular stores, that oscillation frequency is very sensitive to the external Ca2+ concentration and that conversion of the primary oscillation to a propagated Ca2+ wave depends upon IP3-induced Ca2+ release.

Functional innervation of Guinea-pig bladder interstitial cells of cajal subtypes: neurogenic stimulation evokes in situ calcium transients

Several populations of interstitial cells of Cajal (ICC) exist in the bladder, associated with intramural nerves. Although ICC respond to exogenous agonists, there is currently no evidence of their functional innervation. The objective was to determine whether bladder ICC are functionally innervated. Guinea-pig bladder tissues, loaded with fluo-4AM were imaged with fluorescent microscopy and challenged with neurogenic electrical field stimulation (EFS). All subtypes of ICC and smooth muscle cells (SMC) displayed spontaneous Ca2+-oscillations. EFS (0.5Hz, 2Hz, 10Hz) evoked tetrodotoxin (1µM)-sensitive Ca2+-transients in lamina propria ICC (ICC-LP), detrusor ICC and perivascular ICC (PICC) associated with mucosal microvessels. EFS responses in ICC-LP were significantly reduced by atropine or suramin. SMC and vascular SMC (VSM) also responded to EFS. Spontaneous Ca2+-oscillations in individual ICC-LP within networks occurred asynchronously whereas EFS evoked coordinated Ca2+-transients in all ICC-LP within a field of view. Non-correlated Ca2+-oscillations in detrusor ICC and adjacent SMC pre-EFS, contrasted with simultaneous neurogenic Ca2+ transients evoked by EFS. Spontaneous Ca2+-oscillations in PICC were little affected by EFS, whereas large Ca2+-transients were evoked in pre-EFS quiescent PICC. EFS also increased the frequency of VSM Ca2+-oscillations. In conclusion, ICC-LP, detrusor ICC and PICC are functionally innervated. Interestingly, Ca2+-activity within ICC-LP networks and between detrusor ICC and their adjacent SMC were synchronous under neural control. VSM and PICC Ca2+-activity was regulated by bladder nerves. These novel findings demonstrate functional neural control of bladder ICC. Similar studies should now be carried out on neurogenic bladder to elucidate the contribution of impaired nerve-ICC communication to bladder pathophysiology.

Laser-engineered dissolving microneedles for active transdermal delivery of nadroparin calcium

There is an urgent need to replace the injection currently used for low molecular weight heparin (LMWH) multidose therapy with a non- or minimally invasive delivery approach. In this study, laser-engineered dissolving microneedle (DMN) arrays fabricated from aqueous blends of 15% w/w poly(methylvinylether-co-maleic anhydride) were used for the first time in active transdermal delivery of the LMWH nadroparin calcium (NC). Importantly, an array loading of 630 IU of NC was achieved without compromising the array mechanical strength or drug bioactivity. Application of NC-DMNs to dermatomed human skin (DHS) using the single-step 'poke and release' approach allowed permeation of approximately 10.6% of the total NC load over a 48-h study period. The cumulative amount of NC that permeated DHS at 24 h and 48 h attained 12.28 ± 4.23 IU/cm and 164.84 ± 8.47 IU/cm , respectively. Skin permeation of NC could be modulated by controlling the DMN array variables, such as MN length and array density as well as application force to meet various clinical requirements including adjustment for body mass and renal function. NC-loaded DMN offers great potential as a relatively low-cost functional delivery system for enhanced transdermal delivery of LMWH and other macromolecules. © 2012 Elsevier B.V. All rights reserved.

The mechanical effect of the existing cement mantle on the in-cement femoral revision

Cement-in-cement revision hip arthroplasty is an increasingly popular technique to replace a loose femoral stem which retains much of the original cement mantle. However, some concern exists regarding the retention of the existing fatigued and aged cement in such cement-in-cement revisions. This study investigates whether leaving an existing fatigued and aged cement mantle degrades the mechanical performance of a cement-in-cement revision construct.

A Molecular Dynamics Study of the Early Stages of Calcium Carbonate Growth

The precipitation of calcium carbonate in water has been examined using a combination of molecular dynamics and umbrella sampling. During 20 ns molecular dynamics trajectories at elevated calcium carbonate concentrations, amorphous particles are observed to form and appear to be composed of misaligned domains of vaterite and aragonite. The addition of further calcium ions to these clusters is found to be energetically favorable and virtually barrierless. By contrast, there is a large barrier to the addition of calcium to small calcite crystals. Thus, even though calcite nanocrystals are stable in solution, at high supersaturations, particles of amorphous material form because this material grows much faster than ordered calcite nanocrystals.

Binding of Calcium and Carbonate to Polyacrylates

Polyacrylate molecules can be used to slow the growth of calcium carbonate. However, little is known about the mechanism by which the molecules impede the growth rate. A recent computational study (Bulo et al. Macromolecules 2007, 40, 3437) used metadynamics to investigate the binding of calcium to polyacrylate chains and has thrown some light on the coiling and precipitation of these polymers. We extend these simulations to examine the binding of calcium and carbonate to polyacrylate chains. We show that calcium complexed with both carbonate and polyacrylate is a very stable species. The free energies of calcium-carbonate-polyacrylate complexes, with different polymer configurations, are calculated, and differences in the free energy of the binding of carbonate are shown to be due to differences in the amount of steric hindrance about the calcium, which prevents the approach of the carbonate ion.

Ser6 in the maize actin-depolymerizing factor, ZmADF3, is phosphorylated by a calcium-stimulated protein kinase and is essential for the control of functional activity

Maize actin-depolymerizing factor, ZmADF, binds both G- and F-actin and enhances in vitro actin dynamics. Evidence from studies on vertebrate ADF/cofilin supports the view that this class of protein responds to intracellular and extracellular signals and causes actin reorganization. As a test to determine whether such signal-responsive pathways existed in plants, this study addressed the ability of maize ADF to be phosphorylated and the likely effects of such phosphorylation on its capacity to modulate actin dynamics. It is shown that maize ADF3 (ZmADF3) can be phosphorylated by a calcium-stimulated protein kinase present in a 40-70% ammonium sulphate fraction of a plant cell extract. Phosphorylation is shown to be on Ser6, which is only one of nine amino acids that are fully conserved among the ADF/cofilin proteins across distantly related species. In addition, an analogue of phosphorylated ZmADF3 created by mutating Ser6 to Asp6 (zmadf3-4) does not bind G- or F-actin and has little effect on the enhancement of actin dynamics. These results are discussed in context of the previously observed actin reorganization in root hair cells.