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We demonstrated previously that a single injection of recombinant human macrophage colony-stimulating factor (rhM-CSF) is sufficient for osteoclast recruitment and survival in osteopetrotic (op/op) mice with a deficiency in osteoclasts resulting from a mutation in M-CSF gene. In this study, we show that a single injection of recombinant human vascular endothelial growth factor (rhVEGF) can similarly induce osteoclast recruitment in op/op mice. Osteoclasts predominantly expressed VEGF receptor 1 (VEGFR-1), and activity of recombinant human placenta growth factor 1 on osteoclast recruitment was comparable to that of rhVEGF, showing that the VEGF signal is mediated through VEGFR-1. The rhM-CSF–induced osteoclasts died after injections of VEGFR-1/Fc chimeric protein, and its effect was abrogated by concomitant injections of rhM-CSF. Osteoclasts supported by rhM-CSF or endogenous VEGF showed no significant difference in the bone-resorbing activity. op/op mice undergo an age-related resolution of osteopetrosis accompanied by an increase in osteoclast number. Most of the osteoclasts disappeared after injections of anti-VEGF antibody, demonstrating that endogenously produced VEGF is responsible for the appearance of osteoclasts in the mutant mice. In addition, rhVEGF replaced rhM-CSF in the support of in vitro osteoclast differentiation. These results demonstrate that M-CSF and VEGF have overlapping functions in the support of osteoclastic bone resorption.

The evolution of stress in gallium nitride films on sapphire has been measured in real time during metalorganic chemical vapor deposition. In spite of the 16% compressive lattice mismatch of GaN to sapphire, we find that GaN consistently grows in tension at 1050 °C. Furthermore, in situ stress monitoring indicates that there is no measurable relaxation of the tensile growth stress during annealing or thermal cycling.

We have shown recently that by acting on the thyroid-stimulating hormone (TSH) receptor (TSHR), TSH negatively regulates osteoclast differentiation. Both heterozygotic and homozygotic TSHR null mice are osteopenic with evidence of enhanced osteoclast differentiation. Here, we report that the accompanying elevation of TNF␣, an osteoclastogenic cytokine, causes the increased osteoclast differentiation. This enhancement in TSHR ؊/؊ and TSHR ؉/؊ mice is abrogated in compound TSHR ؊͞؊ ͞TNF␣ ؊/؊ and TSHR ؉/؊ ͞ TNF␣ ؉/؊ mice, respectively. In parallel studies, we find that TSH directly inhibits TNF␣ production, reduces the number of TNF␣-producing osteoclast precursors, and attenuates the induction of bone remodeling ͉ osteoclast ͉ macrophage ͉ cytokine A nterior pituitary hormones have long been thought just to stimulate the secretion of master hormones from target endocrine glands, except for our recent demonstration of direct effects of thyroid-stimulating hormone (TSH) and folliclestimulating hormone on the skeleton (1). Thus, until recently, TSH was considered solely to regulate thyroid follicular cell differentiation and thyroid hormone secretion by binding to a seven transmembrane, glycosylated G protein-coupled receptor, the TSH receptor (TSHR). Previous studies had identified TSHRs in other tissues and cells, including the pituitary, thymus, testes, kidney, brain, lymphocytes, adipocytes, and fibroblasts (2, 3), but their functional significance has remained uncertain.Gene ablation studies in mice revealed that TSHR haploinsufficiency did not affect thyroid gland development or function, whereas the total absence of the TSHR expectedly disrupted thyroid follicular structure (4). However, bone mass was reduced not only in homozygote mice but also in the haploinsufficient heterozygotes (5). That TSHR haploinsufficiency, in the absence of a thyroid defect, resulted in osteoporosis established a primary role for the TSHR in bone metabolism. Furthermore, supplementation of TSHR Ϫ/Ϫ mice with thyroid extract to render them euthyroid corrected all hypothyroid abnormalities, including runting, but not reductions in bone mass (5) or sodium-iodide symporter expression (4). The latter observation confirmed that the osteoporosis arose from TSHR deficiency rather than altered thyroid hormone levels. Consistent with this notion, the genetic ablation of the ␣1͞␤ thyroid hormone receptor has been shown not to result in a bone remodeling defect (6).We found that the osteoporosis in TSHR knockout mice was the result of an enhancement in osteoclast differentiation. Consistent with the low bone mass, ex vivo cultures of bone marrow cell precursors from both heterozygote and homozygote mice showed increased osteoclast formation and the enhanced expression of an osteoclast marker tartrate-resistant acid phosphatase (TRAP) (5). This enhanced osteoclast formation was not associated with increased receptor activator of NF-B ligand (RANKL) production but instead with a several-fold increase in the synthesis and release of TNF␣, another...

Abstract. Matrix metalloproteinases (MMPs) play an important role in degeneration of the matrix associated with bone and cartilage. Regulation of osteoclast activity is essential in the treatment of bone disease, including osteoporosis and rheumatoid arthritis. Polyphenols in green tea, particularly epigallocatechin-3-gallate (EGCG), inhibit MMPs expression and activity. However, the effects of the black tea polyphenol, theaflavin-3,3′-digallate (TFDG), on osteoclast and MMP activity are unknown. Therefore, we examined whether TFDG and EGCG affect MMP activity and osteoclast formation and differentiation in vitro. TFDG or EGCG (10 and 100 μM) was added to cultures of rat osteoclast precursors cells and mature osteoclasts. Numbers of multinucleated osteoclasts and actin rings decreased in polyphenol-treated cultures relative to control cultures. MMP-2 and MMP-9 activities were lower in TFDG-and EGCG-treated rat osteoclast precursor cells than in control cultures. MMP-9 mRNA levels declined significantly in TFDG-treated osteoclasts in comparison to control osteoclasts. TFDG and EGCG inhibited the formation and differentiation of osteoclasts via inhibition of MMPs. TFDG may suppress actin ring formation more effectively than EGCG. Thus, TFDG and EGCG may be suitable agents or lead compounds for the treatment of bone resorption diseases.

Objective We evaluated the safety and efficacy of vonoprazan-based amoxicillin and clarithromycin 7-day triple therapy (VAC) in comparison to proton pump inhibitor (PPI)-based (PAC) as a first-line treatment and vonoprazan-based amoxicillin and metronidazole 7-day triple therapy (VAM) in comparison to PPI-based (PAM) as a second-line treatment for the eradication of Helicobacter pylori in Japan. Methods We performed a non-randomized, multi-center, parallel-group study to compare first-line VAC to PAC and second-line VAM to PAM. A pre-planned subgroup analysis on CAM resistance was also performed. Safety was evaluated with an adverse effects questionnaire (AEQ), which was completed by patients during therapy. Results The first-line eradication rates (ER) in the intention-to-treat (ITT) and per protocol (PP) analyses were 84.9% (95% CI: 81.9-87.6%, n=623) and 86.4% (83.5-89.1%, n=612), respectively, for VAC and 78.8% (75.3-82.0%, n=608) and 79.4% (76.0-82.6%, n=603), respectively, for PAC. The ER of VAC was higher than that of PAC in the ITT (p=0.0061) and PP analyses (p=0.0013). The ERs for VAC in patients with CAM-resistant and CAM-susceptible bacteria were 73.2% (59.7-84.2%, n=56) and 88.9% (83.4-93.1%, n=180), respectively. PAC was associated with higher AEQ scores for diarrhea, nausea, headache, and general malaise. In the second-line ITT and PP analyses VAM achieved ERs of 80.5% (74.6-85.6%, n=216) and 82.4% (76.6-87.3%, n=211), respectively, while PAM achieved ERs of 81.5% (74.2-87.4%, n=146) and 82.1% (74.8-87.9%, n=145), respectively. No significant differences were observed in the ITT (p=0.89) or PP (p=1.0) analyses. Conclusion The ER of first-line VAC was higher than that of PAC, but still <90%. No difference was observed between second-line VAM and PAM. Vonoprazan-based triple therapy was safe and well tolerated.

The voltage-gated H ؉ channel is a powerful H ؉ extruding mechanism of osteoclasts, but its functional roles and regulatory mechanisms remain unclear. Electrophysiological recordings revealed that the H ؉ channel operated on activation of protein kinase C together with cell acidosis. Introduction: Hϩ is a key signaling ion in bone resorption. In addition to H ϩ pumps and exchangers, osteoclasts are equipped with H ϩ conductive pathways to compensate rapidly for pH imbalance. The H ϩ channel is distinct in its strong H ϩ extrusion ability and voltage-dependent gatings. Methods: To investigate how and when the H ϩ channel is available in functional osteoclasts, the effects of phorbol 12-myristate 13-acetate (PMA), an activator for protein kinase C, on the H ϩ channel were examined in murine osteoclasts generated in the presence of soluble RANKL (sRANKL) and macrophage-colony stimulating factor (M-CSF). Results and Conclusions:Whole cell recordings clearly showed that the H ϩ current was enhanced by increasing the pH gradient across the plasma membrane (⌬pH), indicating that the H ϩ channel changed its activity by sensing ⌬pH. The reversal potential (V rev ) was a valuable tool for the real-time monitoring of ⌬pH in clamped cells. In the permeabilized patch, PMA (10 nM-1.6 M) increased the current density and the activation rate, slowed decay of tail currents, and shifted the threshold toward more negative voltages. In addition, PMA caused a negative shift of V rev , suggesting that intracellular acidification occurred. The PMA-induced cell acidosis was confirmed using a fluorescent pH indicator (BCECF), which recovered quickly in a K

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