Sodium/hydrogen exchanger NHA2 in osteoclasts: subcellular localization and role in vitro and in vivo

NHA2 was recently identified as a novel sodium/hydrogen exchanger which is strongly upregulated during RANKL-induced osteoclast differentiation. Previous in vitro studies suggested that NHA2 is a mitochondrial transporter required for osteoclast differentiation and bone resorption. Due to the lack of suitable antibodies, NHA2 was studied only on RNA level thus far. To define the protein's role in osteoclasts in vitro and in vivo, we generated NHA2-deficient mice and raised several specific NHA2 antibodies. By confocal microscopy and subcellular fractionation studies, NHA2 was found to co-localize with the late endosomal and lysosomal marker LAMP1 and the V-ATPase a3 subunit, but not with mitochondrial markers. Immunofluorescence studies and surface biotinylation experiments further revealed that NHA2 was highly enriched in the plasma membrane of osteoclasts, localizing to the basolateral membrane of polarized osteoclasts. Despite strong upregulation of NHA2 during RANKL-induced osteoclast differentiation, however, structural parameters of bone, quantified by high-resolution microcomputed tomography, were not different in NHA2-deficient mice compared to wild-type littermates. In addition, in vitro RANKL stimulation of bone marrow cells isolated from wild-type and NHA2-deficient mice yielded no differences in osteoclast development and activity. Taken together, we show that NHA2 is a RANKL-induced plasmalemmal sodium/hydrogen exchanger in osteoclasts. However, our data from NHA2-deficient mice suggest that NHA2 is dispensable for osteoclast differentiation and bone resorption both in vitro and in vivo.

Sodium/hydrogen exchanger NHA2 is critical for insulin secretion in β-cells

NHA2 is a sodium/hydrogen exchanger with unknown physiological function. Here we show that NHA2 is present in rodent and human β-cells, as well as β-cell lines. In vivo, two different strains of NHA2-deficient mice displayed a pathological glucose tolerance with impaired insulin secretion but normal peripheral insulin sensitivity. In vitro, islets of NHA2-deficient and heterozygous mice, NHA2-depleted Min6 cells, or islets treated with an NHA2 inhibitor exhibited reduced sulfonylurea- and secretagogue-induced insulin secretion. The secretory deficit could be rescued by overexpression of a wild-type, but not a functionally dead, NHA2 transporter. NHA2 deficiency did not affect insulin synthesis or maturation and had no impact on basal or glucose-induced intracellular Ca(2+) homeostasis in islets. Subcellular fractionation and imaging studies demonstrated that NHA2 resides in transferrin-positive endosomes and synaptic-like microvesicles but not in insulin-containing large dense core vesicles in β-cells. Loss of NHA2 inhibited clathrin-dependent, but not clathrin-independent, endocytosis in Min6 and primary β-cells, suggesting defective endo-exocytosis coupling as the underlying mechanism for the secretory deficit. Collectively, our in vitro and in vivo studies reveal the sodium/proton exchanger NHA2 as a critical player for insulin secretion in the β-cell. In addition, our study sheds light on the biological function of a member of this recently cloned family of transporters.

SOLAR RADIATION PRESSURE AND LOCAL INTERSTELLAR MEDIUM FLOW PARAMETERS FROM INTERSTELLAR BOUNDARY EXPLORER LOW ENERGY HYDROGEN MEASUREMENTS

Neutral hydrogen atoms that travel into the heliosphere from the local interstellar medium (LISM) experience strong effects due to charge exchange and radiation pressure from resonant absorption and re-emission of Lyα. The radiation pressure roughly compensates for the solar gravity. As a result, interstellar hydrogen atoms move along trajectories that are quite different than those of heavier interstellar species such as helium and oxygen, which experience relatively weak radiation pressure. Charge exchange leads to the loss of primary neutrals from the LISM and the addition of new secondary neutrals from the heliosheath. IBEX observations show clear effects of radiation pressure in a large longitudinal shift in the peak of interstellar hydrogen compared with that of interstellar helium. Here, we compare results from the Lee et al. interstellar neutral model with IBEX-Lo hydrogen observations to describe the distribution of hydrogen near 1 AU and provide new estimates of the solar radiation pressure. We find over the period analyzed from 2009 to 2011 that radiation pressure divided by the gravitational force (μ) has increased slightly from μ = 0.94 ± 0.04 in 2009 to μ = 1.01 ± 0.05 in 2011. We have also derived the speed, temperature, source longitude, and latitude of the neutral H atoms and find that these parameters are roughly consistent with those of interstellar He, particularly when considering the filtration effects that act on H in the outer heliosheath. Thus, our analysis shows that over the period from 2009 to 2011, we observe signatures of neutral H consistent with the primary distribution of atoms from the LISM and a radiation pressure that increases in the early rise of solar activity.

Enamel Surface Changes After Exposure to Bleaching Gels Containing Carbamide Peroxide or Hydrogen Peroxide.

OBJECTIVE This study evaluated the differences in enamel color change, surface hardness, elastic modulus, and surface roughness between treatments with four bleaching gels containing carbamide peroxide (two at 10% and one each at 35%, and 45%) and two bleaching gels containing hydrogen peroxide (two at 40%). METHODS Enamel specimens were bleached and color changes were measured. Color change was calculated using either ΔE or the Bleaching Index (BI). Then, surface hardness, elastic modulus, and surface roughness of the enamel specimens were evaluated. All measurements were performed at baseline and directly after the first bleaching treatment for all carbamide peroxide- and hydrogen peroxide-containing bleaching gels. In addition, final measurements were made 24 hours after each of a total of 10 bleaching treatments for carbamide peroxide bleaching gels, and 1 week after each of a total of three bleaching treatments for hydrogen peroxide bleaching gels. RESULTS After the last bleaching treatment, respective ΔE scores were 17.6 and 8.2 for the two 10% carbamide peroxide gels, 12.9 and 5.6 for the 45% and 35% carbamide peroxide gels, and 9.6 and 13.9 for the two 40% hydrogen peroxide gels. The respective BI scores were -2.0 and -2.0 for the two 10% carbamide peroxide gels, -3.5 and -1.5 for the 45% and 35% carbamide peroxide gels, and -2.0 and -3.0 for the two 40% hydrogen peroxide gels. Each bleaching gel treatment resulted in significant whitening; however, no significant difference was found among the gels after the last bleaching. Whitening occurred within the first bleaching treatments and did not increase significantly during the remaining treatments. Surface hardness significantly decreased after the last bleaching treatment, when 10% carbamide peroxide was used. Furthermore, significant changes in the elastic modulus or surface roughness occurred only after treatment with 10% carbamide peroxide. CONCLUSION All six bleaching gels effectively bleached the enamel specimens independent of their concentration of peroxide. Gels with low peroxide concentration and longer contact time negatively affected the enamel surface.