Epitaxial growth of FeSe0.5_{0.5}Te0.5_{0.5} thin films on CaF2_2 substrates with high critical current density

In-situ epitaxial growth of FeSe$_{0.5}$Te$_{0.5}$ thin films is demonstrated on a non-oxide substrate CaF$_2$. Structural analysis reveals that compressive stress is moderately added to 36-nm thick FeSe$_{0.5}$Te$_{0.5}$, which pushes up the critical temperature above 15 K, showing higher values than that of bulk crystals. Critical current density at $T$ = 4.5 K reaches 5.9 x 10$^4$ Acm$^{-2}$ at $\mu_0H$ = 10 T, and 4.2 x 10$^4$ Acm$^{-2}$ at $\mu_0H$ = 14 T. These results indicate that fluoride substrates have high potential for the growth of iron-based superconductors in comparison with popular oxide substrates.Comment: 9 pages, 3 figures, to be published in Applied Physics Express 4, 053101 (2011

Bone morphogenetic protein-2 (BMP-2) and transforming growth factor-β1 (TGF-β1) alter connexin 43 phosphorylation in MC3T3-E1 Cells

BACKGROUND: Bone morphogenetic proteins (BMPs) and transforming growth factor-βs (TGF-βs) are important regulators of bone repair and regeneration. BMP-2 and TGF-β1 have been shown to inhibit gap junctional intercellular communication (GJIC) in MC3T3-E1 cells. Connexin 43 (Cx43) has been shown to mediate GJIC in osteoblasts and it is the predominant gap junctional protein expressed in these murine osteoblast-like cells. We examined the expression, phosphorylation, and subcellular localization of Cx43 after treatment with BMP-2 or TGF-β1 to investigate a possible mechanism for the inhibition of GJIC. RESULTS: Northern blot analysis revealed no detectable change in the expression of Cx43 mRNA. Western blot analysis demonstrated no significant change in the expression of total Cx43 protein. However, significantly higher ratios of unphosphorylated vs. phosphorylated forms of Cx43 were detected after BMP-2 or TGF-β1 treatment. Immunofluorescence and cell protein fractionation revealed no detectable change in the localization of Cx43 between the cytosol and plasma membrane. CONCLUSIONS: BMP-2 and TGF-β1 do not alter expression of Cx43 at the mRNA or protein level. BMP-2 and TGF-β1 may inhibit GJIC by decreasing the phosphorylated form of Cx43 in MC3T3-E1 cells

A new technique for precisely and accurately measuring lumbar spine bone mineral density in mice using clinical dual energy X-ray absorptiometry (DXA)

Dual Energy X-ray Absorptiometry (DXA) is effective in measuring bone mineral density (BMD) in mice for early detection of osteoporosis. However, scanners designed for use with small animals (i.e. PIXImus) are very expensive. Used human DXA machines are cheaper to obtain, but analysis of scans from these instruments is operator-dependent. Obtaining reliable data depends on having a single operator analyze the scans in a blinded fashion. Scan quality is improved by excising the bone prior to scanning, which does not allow serial measurements. This study describes a novel method of analyzing lumbar spine BMD in mice using whole body DXA. This non-invasive technique has a high degree of precision and reproducibility, with good correlation between multiple observers. Inter-observer variability (0.063 ± 0.00317 g/cm2 [mean ± SD], 5.05 [% coefficient of variation (CV)], repeat scan variability (0.063 ± 0.00364 g/cm2 [mean ± SD], 5.94 [%CV]) were very low compared to variability between different animals (0.063 ± 0.00588 g/cm2 [mean ± SD], 9.64 [%CV]) and variability seen in same animal over time (0.011 ± 0.00885 g/cm2 [mean ± SD], 80.68 [%CV]). The measurement error is thus smaller than the biological variation. Accuracy was determined by comparing average peak BMD from two scans per mouse in-vivo (0.066 g/cm2) versus excised spine (0.065 g/cm2). Furthermore, correlation between bone ash weights and whole body lumbar spine BMD measurements (p < 0.0001) was highly significant. This technique thus shows a high degree of precision and accuracy, even with multiple observers, for measuring BMD in mice using a DXA machine designed for clinical use

The second intracellular loop of the calcitonin gene-related peptide receptor provides molecular determinants for signal transduction and cell surface expression

The calcitonin gene-related peptide (CGRP) receptor is a heterodimer of a family B G-protein-coupled receptor, calcitonin receptor-like receptor (CLR), and the accessory protein receptor activity modifying protein 1. It couples to Gs, but it is not known which intracellular loops mediate this. We have identified the boundaries of this loop based on the relative position and length of the juxtamembrane transmembrane regions 3 and 4. The loop has been analyzed by systematic mutagenesis of all residues to alanine, measuring cAMP accumulation, CGRP affinity, and receptor expression. Unlike rhodopsin, ICL2 of the CGRP receptor plays a part in the conformational switch after agonist interaction. His-216 and Lys-227 were essential for a functional CGRP-induced cAMP response. The effect of (H216A)CLR is due to a disruption to the cell surface transport or surface stability of the mutant receptor. In contrast, (K227A)CLR had wild-type expression and agonist affinity, suggesting a direct disruption to the downstream signal transduction mechanism of the CGRP receptor. Modeling suggests that the loop undergoes a significant shift in position during receptor activation, exposing a potential G-protein binding pocket. Lys-227 changes position to point into the pocket, potentially allowing it to interact with bound G-proteins. His-216 occupies a position similar to that of Tyr-136 in bovine rhodopsin, part of the DRY motif of the latter receptor. This is the first comprehensive analysis of an entire intracellular loop within the calcitonin family of G-protein-coupled receptor. These data help to define the structural and functional characteristics of the CGRP-receptor and of family B G-protein-coupled receptors in general. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc

The basic helix loop helix transcription factor twist1 is a novel regulator of ATF4 in osteoblasts

Parathyroid hormone (PTH) is an essential regulator of endochondral bone formation and an important anabolic agent for the reversal of bone loss. PTH mediates its functions in part by regulating binding of the bone‐related activating transcription factor 4 (ATF4) to the osteoblast‐specific gene, osteocalcin. The basic helix‐loop‐helix (bHLH) factors Twist1 and Twist2 also regulate osteocalcin transcription in part through the interaction of the C‐terminal “box” domain in these factors and Runx2. In this study, we discovered a novel function of PTH: its ability to dramatically decrease Twist1 transcription. Since ATF4 is a major regulator of the PTH response in osteoblasts, we assessed the mutual regulation between these factors and determined that Twist proteins and ATF4 physically interact in a manner that affects ATF4 DNA binding function. We mapped the interaction domain of Twist proteins to the C‐terminal “box” domain and of ATF4, to the N‐terminus. Furthermore, we demonstrate that Twist1 overexpression in osteoblasts attenuates ATF4 binding to the osteocalcin promoter in response to PTH. This study thus identifies Twist proteins as novel inhibitory binding partners of ATF4 and explores the functional significance of this interaction. J. Cell. Biochem. 113: 70–79, 2012. © 2011 Wiley Periodicals, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/89496/1/23329_ftp.pd

Immunocompetent murine model of cancer cachexia for head and neck squamous cell carcinoma

BackgroundMuscle wasting and weight loss were observed when carcinomas were induced in a murine model of head and neck squamous cell carcinomas. Our hypothesis was C3H/HeN mice would develop evidence of cachexia when injected with tumor cellsMethodsAge- and weight-matched female mice were injected with SCCF/VII cells. Daily food intake and weights were measured. Body composition and analysis of circulating cytokines was performed. At the completion of experiments, hind legs were weighed. Muscle atrophy was detected using analysis for muscle ring finger 1 (MuRF1). The tumor-derived lipid mobilizing factor (LMF) was measured.ResultsDespite increased food intake, tumor-bearing mice lost weight and experienced reduced hind leg weights. Interleukin-1beta (IL-1beta) was increased and MuRF1 was present in tumor-bearing mice but not controls. LMF was present in SCCF/VII cells.ConclusionIn this immunocompetent murine model, we demonstrated the development of cancer cachexia in mice inoculated with SCCF cells, which express LMF. There was increased serum IL-1beta, weight loss, and muscle wasting and atrophy in tumor-bearing mice

PTH Receptor Signaling in Osteocytes Governs Periosteal Bone Formation and Intracortical Remodeling

The periosteal and endocortical surfaces of cortical bone dictate the geometry and overall mechanical properties of bone. Yet the cellular and molecular mechanisms that regulate activity on these surfaces are far from being understood. Parathyroid hormone (PTH) has profound effects in cortical bone, stimulating periosteal expansion and at the same time accelerating intracortical bone remodeling. We report herein that transgenic mice expressing a constitutive active PTH receptor in osteocytes (DMP1-caPTHR1 mice) exhibit increased cortical bone area and an elevated rate of periosteal and endocortical bone formation. In addition, DMP1-caPTHR1 mice display a marked increase in intracortical remodeling and cortical porosity. Crossing DMP1-caPTHR1 mice with mice lacking the Wnt coreceptor, LDL-related receptor 5 (LRP5), or with mice overexpressing the Wnt antagonist Sost in osteocytes (DMP1-Sost mice) reduced or abolished, respectively, the increased cortical bone area, periosteal bone formation rate, and expression of osteoblast markers and Wnt target genes exhibited by the DMP1-caPTHR1 mice. In addition, DMP1-caPTHR1 lacking LRP5 or double transgenic DMP1-caPTHR1;DMP1-Sost mice exhibit exacerbated intracortical remodeling and increased osteoclast numbers, and markedly decreased expression of the RANK decoy receptor osteoprotegerin. Thus, whereas Sost downregulation and the consequent Wnt activation is required for the stimulatory effect of PTH receptor signaling on periosteal bone formation, the Wnt-independent increase in osteoclastogenesis induced by PTH receptor activation in osteocytes overrides the effect on Sost. These findings demonstrate that PTH receptor signaling influences cortical bone through actions on osteocytes and defines the role of Wnt signaling in PTH receptor action. © 2011 American Society for Bone and Mineral Research

PTH Signaling During Exercise Contributes to Bone Adaptation

Improving the structural integrity of bone reduces fracture risk and development of osteoporosis later in life. Exercise can increase the mechanical properties of bone, and this increase is often attributed to the dynamic loading created during exercise. However, the increase in systemic parathyroid hormone (PTH) levels during exercise gives reason to hypothesize that PTH signaling also regulates bone adaptation in response to exercise. Therefore, the first aim of this study was to establish the impact PTH signaling has on bone adaptation during exercise by inhibiting PTH signaling with PTH(7‐34); the second aim was to determine whether increasing PTH levels during exercise with PTH(1‐34) can augment bone adaptation. Thirty minutes after a single bout of running on a treadmill, mice exhibited a twofold increase in systemic PTH levels. Under the same exercise regimen, the influence of PTH signaling on bone adaptation during exercise was then evaluated in mice after 21 consecutive days of exercise and treatment with PTH(7‐34), PTH(1‐34), or vehicle. Exercise alone caused a significant increase in trabecular bone volume with adaptation to a more platelike structure, which was inhibited with PTH(7‐34) during exercise. Changes in structural‐level and tissue‐level mechanical properties during exercise occurred in the absence of significant changes to cortical bone geometry. Inhibition of PTH signaling during exercise attenuated the changes in structural‐level mechanical properties, but not tissue‐level properties. Enhanced PTH signaling during exercise with PTH(1‐34) increased trabecular and cortical bone volume, but had little effect on the structural‐level and tissue‐level mechanical properties compared to exercise alone. Our study is the first to demonstrate that bone adaptation during exercise is not only a function of dynamic loading, but also PTH release, and that PTH signaling contributes differently at the structural and tissue levels. © 2015 American Society for Bone and Mineral Research.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/111785/1/jbmr2432.pd