CaSR-Mediated hBMSCs Activity Modulation: Additional Coupling Mechanism in Bone Remodeling Compartment

Near the bone remodeling compartments (BRC), extracellular calcium concentration (Ca2+o) is locally elevated and bone marrow stromal cells (BMSCs) close to the BRC can be exposed to high calcium concentration. The calcium-sensing receptor (CaSR) is known to play a key role in maintaining extracellular calcium homeostasis by sensing fluctuations in the levels of extracellular calcium (Ca2+o). When human BMSCs (hBMSCs) were exposed to various calcium concentrations (1.8, 3, 5, 10, 30 mM), moderate-high extracellular calcium concentrations (3–5 mM) stimulated proliferation, while a high calcium concentration (30 mM) inhibited the proliferation. Exposure to various calcium concentrations did not induce significant differences in the apoptotic cell fraction. Evaluation of multi-lineage differentiation potential showed no significant difference among various calcium concentration groups, except for the high calcium concentration (30 mM) treated group, which resulted in increased calcification after in vitro osteogenic differentiation. Treatment of NPS2143, a CaSR inhibitor, abolished the stimulatory effect on hBMSCs proliferation and migration indicating that CaSR is involved. These results suggest that the calcium concentration gradient near the BRC may play an important role in bone remodeling by acting as an osteoblast–osteoclast coupling mechanism through CaSR.

The Photoconvertible Fluorescent Probe, CaMPARI, Labels Active Neurons in Freely-Moving Intact Adult Fruit Flies

Linking neural circuitry to behavior by mapping active neurons in vivo is a challenge. Both genetically encoded calcium indicators (GECIs) and intermediate early genes (IEGs) have been used to pinpoint active neurons during a stimulus or a behavior but have drawbacks such as limiting the movement of the organism, requiring a priori knowledge of the active region or having poor temporal resolution. CaMPARI (calcium-modulated photoactivatable ratiometric integrator) was engineered to overcome these spatial-temporal challenges. CaMPARI is a photoconvertible protein that only converts from green to red fluorescence in the presence of high calcium concentration and violet (405 nm) light. This allows the experimenter to precisely mark active neurons within defined temporal windows. The photoconversion can then be quantified by taking the ratio of the red fluorescence to the green. CaMPARI promises the ability to trace active neurons during specific stimulus; however, CaMPARI’s uses in adult Drosophila have been limited to photoconversion during fly immobilization. Here, we demonstrate a method that allows photoconversion of multiple freely-moving intact adult flies during a stimulus. Flies were placed in a dish with filter paper wet with acetic acid (pH=2) or neutralized acetic acid (pH=7) and exposed to photoconvertible light (60 mW) for 30 min (500 ms on, 200 ms off). Immediately following photoconversion, whole flies were fixed and imaged by confocal microscopy. The red:green ratio was quantified for the DC4 glomerulus, a bundle of neurons expressing Ir64a, an ionotropic receptor that senses acids in the Drosophila antennal lobe. Flies exposed to acetic acid showed 1.3-fold greater photoconversion than flies exposed to neutralized acetic acid. This finding was recapitulated using a more physiological stimulus of apple cider vinegar. These results indicate that CaMPARI can be used to label neurons in intact, freely-moving adult flies and will be useful for identifying the circuitry underlying complex behaviors.

Construction and Demolition Waste as Recycled Aggregates in Alkali-Activated Concretes

The growth of global construction has contributed to an inevitable increase in the amount of construction and demolition (C&D) waste, and the recycling of C&D waste as aggregates in concrete is receiving increased interest, resulting in less demand for normal aggregates and bringing a potential solution for the landfilling of wastes. Recently, several studies have focused on the use of C&D waste in alkali-activated concrete to move one step closer to sustainable concretes. This paper focuses on the main mechanisms of using C&D waste in the resulting physical, mechanical, and durability properties of alkali-activated concrete in fresh and hardened state properties. The main difficulties observed with recycled aggregates (RA) in concrete, such as high levels of water demand, porous structure, and low mechanical strength, occur in RA alkali-activated concretes. These are associated with the highly porous nature and defects of RA. However, the high calcium concentration of RA affects the binder gel products, accelerates the hardening rate of the concrete, and reduces the flowability of alkali-activated concretes. For this reason, several techniques have been investigated for modifying the water content and workability of the fresh matrix and for treating RA and RA/alkali-activated binder interactions to produce more sustainable alkali-activated concretes.

Interactions between calmodulin and neurogranin govern the dynamics of CaMKII as a leaky integrator

Calmodulin-dependent kinase II (CaMKII) has long been known to play an important role in learning and memory as well as long term potentiation (LTP). More recently it has been suggested that it might be involved in the time averaging of synaptic signals, which can then lead to the high precision of information stored at a single synapse. However, the role of the scaffolding molecule, neurogranin (Ng), in governing the dynamics of CaMKII is not yet fully understood. In this work, we adopt a rule-based modeling approach through the Monte Carlo method to study the effect of Ca2+ signals on the dynamics of CaMKII phosphorylation in the postsynaptic density (PSD). Calcium surges are observed in synaptic spines during an EPSP and back-propagating action potential due to the opening of NMDA receptors and voltage dependent calcium channels. We study the differences between the dynamics of phosphorylation of CaMKII monomers and dodecameric holoenzymes. The scaffolding molecule Ng, when present in significant concentration, limits the availability of free calmodulin (CaM), the protein which activates CaMKII in the presence of calcium. We show that it plays an important modulatory role in CaMKII phosphorylation following a surge of high calcium concentration. We find a non-intuitive dependence of this effect on CaM concentration that results from the different affinities of CaM for CaMKII depending on the number of calcium ions bound to the former. It has been shown previously that in the absence of phosphatase CaMKII monomers integrate over Ca2+ signals of certain frequencies through autophosphorylation (Pepke et al, Plos Comp. Bio., 2010). We also study the effect of multiple calcium spikes on CaMKII holoenzyme autophosphorylation, and show that in the presence of phosphatase CaMKII behaves as a leaky integrator of calcium signals, a result that has been recently observed in vivo. Our models predict that the parameters of this leaky integrator are finely tuned through the interactions of Ng, CaM, CaMKII, and PP1. This is a possible mechanism to precisely control the sensitivity of synapses to calcium signals.

Hard water syndrome: a case series of 30 patients from a London haemodialysis unit

Severe and life-threatening hypercalcaemia can develop in haemodialysis patients dialysed against a dialysate with a high calcium concentration, the so-called hard water syndrome. Here we describe the development of hard water syndrome in 30 patients following sequential failure of the reverse osmosis unit and water softeners. Serum calcium levels rose from 2.43 ± 0.19 to 3.92 ± 0.51 mmol/L after exposure. All patients required emergency haemodialysis and four acutely deteriorated, one of whom was 24 weeks pregnant. This is the largest reported series of patients affected by this rare and severe condition. This event led to the introduction of processes to minimize future risks.

Inhibitory effect of high calcium concentration on municipal solid waste leachate treatment by the activated sludge process

This research focused on the inhibitory effects of Ca on the aerobic biological treatment of landfill leachate containing extremely high Ca concentrations. When the Ca concentration in leachate to be treated was more than 4500 mg l−1, the total organic carbon removal rate was significantly reduced and the processing time to achieve the same removal efficiency was 1.4 times that in the control treatment without added Ca. In contrast, the total nitrogen and ammonia nitrogen (NH4+–N) removal efficiencies were positively related to the Ca concentration, increasing from 65.2% to 81.2% and from 69.2% to 83.7%, respectively, when the dosage of added Ca increased from zero to 8000 mg l−1. During aerobic treatment, the reductions of solution Ca concentration were in the range of 1003–2274 mg l−1 and were matched with increases in the Ca content in the residual sludge. The inhibition threshold of Ca in the leachate treated by the activated sludge process appeared to be 4500 mg l−1, which could be realized by controlling the influent Ca concentration and using an appropriate sludge return ratio in the activated sludge process.

Deficiencies in extrusion of the second polar body due to high calcium concentrations during in vitro fertilization in inbred C3H/He mice

SummarySuccessful in vitro fertilization (IVF) of all inbred strains of laboratory mice has not yet been accomplished. We have previously shown that a high calcium concentration improved IVF in various inbred mice. However, we also found that in cumulus-free ova of C3H/He mice such IVF conditions significantly increased the deficiency of extrusion of the second polar body (PBII) in a dose-dependent manner (2% at 1.71 mM and 29% at 6.84 mM, P < 0.05) and that PBII extrusion was affected by high calcium levels at 2–3 h post-insemination. While developmental competence of ova without PBII extrusion to blastocysts after 96 h culture was not affected, a significant reduction in the nuclear number of the inner cell mass was observed in blastocyst fertilized under high calcium condition. We also examined how high calcium concentration during IVF affects PBII extrusion in C3H/He mice. Cumulus cells cultured under high calcium conditions showed a significantly alleviated deficient PBII extrusion. This phenomenon is likely to be specific to C3H/He ova because deficient PBII extrusion in reciprocal fertilization between C3H and BDF1 gametes was observed only in C3H/He ova. Sperm factor(s) was still involved in deficient PBII extrusion due to high calcium concentrations, as this phenomenon was not observed in ova activated by ethanol. The cytoskeletal organization of ova without PBII extrusion showed disturbed spindle rotation, incomplete formation of contractile ring and disturbed localization of actin, suggesting that high calcium levels affect the anchoring machinery of the meiotic spindle. These results indicate that in C3H/He mice high calcium levels induce abnormal fertilization, i.e. deficient PBII extrusion by affecting the cytoskeletal organization, resulting in disturbed cytokinesis during the second meiotic division. Thus, use of high calcium media for IVF should be avoided for this strain.