bluuebunny/crossref_metadata_2025 huggingface dataset:
Photosensitized Two-electron Reduction of Viologen and the Reactivity of the Neutral Viologen
Authors: Park Joon Woo , Kim Jung Ha , Hwang Book Kee , Park Kwanghee Koh
Published: April 2006

The initially formed viologen radical cation (V•+) is further reduced to yield neutral viologen (V°) by Ru(bpy)32+ sensitized photochemical reduction. By the reactions with water, 75–80% of V° is reoxidized to V+ and the remainder is destroyed. The decreasing order of the reactivity is V° in the absence of β-cyclodextrin (β-CD) > V° in the presence of β-CD > V° covalently bound to β-CD.
Synthesis of 3-(1-Adamantyl)pyrazole and 3,5-Di(1-adamantyl)pyrazole in a Microwave Oven
Authors: Forfar Isabelle , Cabildo Pilar , Claramunt Rosa María , Elguero José
Published: April 2006

An extraordinary inversion of the regioselectivity in the reaction of C-adamantylation of pyrazoles by 1-bromoadamantane was observed when the autoclave vessel was replaced by a microwave oven. In these last conditions mono and disubstituted 3(5)-(1-adamantyl) pyrazoles were obtained.
Remarkable Salt Effects in the Highly Enhanced Enantioselective Hydrolysis of Amino Acid Esters with the Active Tripeptide in the Vesicular System
Authors: Goto Koichi , Imamura Chikara , Yamamoto Shinichi , Matsumoto Yoko , Ueoka Ryuichi
Published: April 2006

The remarkably high enantioselectivity (kLa,obsd/kDa,obsd = 67) was attained along with the large rate-enhancement of the L-form substrate for the hydrolytic cleavage of N-dodecanoyl-D(L)-Phe-PNP by controlling the ionic strength ([KCl] = 0.03 M, 1 M = 1 mol dm−3) in the vesicular system.
An FTIR Study of the Species Adsorbed on a Rh/Al2O3 Catalyst in the Selective Reduction of NO by Propylene
Authors: Bamwenda Gratian R , Obuchi Akira , Ogata Atsushi , Mizuno Koichi
Published: April 2006

An in situ FTIR study of the reduction of NO by propylene in O2 has shown that the reaction on the Rh/Al2O3 surface at 623 K leads to the formation of IR bands assigned to adsorbed hydrocarbons, -NCO, -CN, -NO+, -CO32− and CO2 species. The intensity of the bands depends on the reaction time and the presence or absence of the individual reactants.
Time-resolved Infrared Spectroscopic Diagnosis of a Bimolecular Chemical Reaction in Room Temperature Solution. The CCl3 Radical Produced from Photoexcited trans-Stilbene and Carbon Tetrachloride
Authors: Iwata Koichi , Kataoka Shuichi , Hamaguchi Hiro-o
Published: April 2006

The newly found photochemical reaction between trans-stilbene and carbon tetrachloride was examined with time-resolved infrared spectroscopy. On the photoexcitation of trans-stilbene in a carbon tetrachloride solution at room temperature, an intense infrared band was observed at 896 cm−1. This band is identified as a band from the trichloromethyl (CCl3) radical. It has been clarified that photoexcited trans-stilbene abstracts hydrogen from carbon tetrachloride to generate the CCl3 radical.
An ab Initio MO Study on Gas-Phase Adsorptive Interactions of Aromatic Compounds onto the Surface of Silica Gel
Authors: Suzuki Tetsuo , Tamon Hajime , Okazaki Morio
Published: April 2006

Microscopic features of gas-phase adsorption phenomena of a few aromatic compounds on the surface of silica gel have been investigated by ab initio molecular orbital (MO) calculations. The interaction energies and the structures of the adsorbate-adsorbent systems have been discussed. It has been found that interactions between the substituent groups of the aromatic compounds and silanol groups of silica gel contribute much to the adsorption.
High-Pressure Promoted and Silica Gel Catalyzed Aminolysis of Epoxides with Glycine Esters
Authors: Kotsuki Hiyoshizo , Shimanouchi Tomoyasu , Teraguchi Masahiro , Kataoka Masanori , Tatsukawa Akira , Nishizawa Hitoshi
Published: April 2006

A variety of epoxides are efficiently reacted with tert-butyl glycinates under high pressure conditions or by supporting on silica gel to afford the corresponding N-(β-hydroxyalkyl)glycine esters.
Oxalate Formation in Electrochemical CO2 Reduction Catalyzed by Rhodium-Sulfur Cluster
Authors: Kushi Yoshinori , Nagao Hirotaka , Nishioka Takanori , Isobe Kiyoshi , Tanaka Koji
Published: April 2006

Electrochemical reduction of CO2 catalyzed by a triangular rhodium complex [(RhCp*)3(μ3-S)2]2+ selectively produced formate and oxalate in the presence of Bu4NBF4 and LiBF4, respectively, under the controlled potential electrolysis at −1.50 V (vs. SCE) in CO2-saturated CH3CN. A solution IR spectrum evidenced the adduct formation between [(RhCp*)3(μ3-S)2]0 and CO2 as the possible precursor for the oxalate formation.
Photoluminescence of Layered Potassium Niobates (K4Nb6O17 and KNb3O8) and Effects of Hydration and H+-exchange on the Luminescence Properties
Authors: Kudo Akihiko , Sakata Tadayoshi
Published: April 2006

KNb3O8 showed blue luminescence of 450 nm at 300 K. The emission spectra of K4Nb6O17 and KNb3O8 at 77 K were red-shifted by H2O intercalation and H+-exchange, indicating that the photochemical properties of these hosts were changed with interlayer conditions.
Novel Preparation of Polyaniline Film from Poly(anthranilic acid) as a Precursor
Authors: Toshima Naoki , Yan Hu , Gotoh Yaeko , Ishiwatari Mariko
Published: April 2006

Poly(anthranilic acid), prepared by oxidative polymerization of anthranilic acid with ammonium peroxodisulfate, is soluble in N-methyl-2-pyrrolidone. The casting of the resulting solution forms a smooth film of poly(anthranilic acid). Heat treatment of the poly(anthranilic acid) film at about 240 °C produces a polyaniline film accompanying the elimination of carbon dioxide, which can provide a new method of preparation of a polyaniline film from the precursor.
Essential Role of Acidity in the Catalytic Reduction of Nitrogen Monoxide by Methane in the Presence of Oxygen over Palladium-Loaded Zeolites
Authors: Nishizaka Yukiko , Misono Makoto
Published: April 2006

It has been demonstrated that the protonic acidity is essential for the reduction of nitrogen monoxide by methane over Pd ion-exchanged H-ZSM-5 (Pd/HZ), by the comparison of the reaction with that over Pd ion-exchanged Na-ZSM-5 (Pd/NaZ), having similar dispersion of palladium. It was further shown that the difference between the active Pd/HZ and the inactive Pd/NaZ is present in the reaction between NO2 and CH4.
Selective Dimerization and Hydrogenation of Ethylene on Active Zirconium Oxide with Coordinatively Unsaturated Surface Sites
Authors: Moon Sang-Chul , Tsuji Kouji , Nomura Takaiki , Anpo Masakazu
Published: April 2006

The evacuation of the ZrO2 catalyst at higher temperatures led to the appearance of an abnormal absorption and photoluminescence due to the formation of coordinatively unsaturated surface sites. These active ZrO2 catalysts having coordinatively unsaturated surface sites exhibited a high catalytic activity for the selective dimerization of C2H4 to 2-C4H8. In the presence of H2 and C2H4, the formation of C4H10 was observed in place of 2-C4H8. The significant role of the coordinatively unsaturated surface sites in these reactions on the active ZrO2 catalyst has been clarified in detail for the first time.
Novel Generation, Characterization, and Trapping of 2-Methylene-3-cyclobutene-1-selones
Authors: Shimada Kazuaki , Akimoto Satoru , Takikawa Yuji , Kabuto Chizuko
Published: April 2006

Heating of alkynyl propargyl selenides afforded a stereoisomeric mixture of 5,10-diselenadispiro[3.1.3.1]deca-1,7-dienes. The 1,3-diselenetane rings of the products caused cycloreversion by heating to generate 2-methylene-3-cyclobutene-1-selones and the selones were trapped with diene to give [4+2] cycloadducts.
An Improved Purification Method for the Rapid Synthesis of High Purity Fluorobutyrophenone Neuroleptics from Nitro and Chloro Precursors Suitable for PET Study
Authors: Hashizume Kazunari , Hashimoto Naoto , Cork David G , Miyake Yoshihiro
Published: April 2006

A rapid micro-scale synthesis of fluorinated neuroleptics with extremely high chemical purity was accomplished using a new single column reverse-phase HPLC purification procedure that employs a strongly alkaline eluent to clearly separate F-labeled compounds from the large excess of nitro or chloro precursor. The method is applicable to the production of 18F-labeled positron emission tomography (PET) tracers with high specific activity.
Precise Discrimination between Butyl and Pentyl Groups in Molecular Aggregates
Authors: Endo Tadashi , Isago Takashi , Gohara Hideshi , Watanabe Susumu , Yazawa Takashi
Published: April 2006

In oxidation of a pair of associating thiols, one with a straight-chain alkyl or a cyclic hydrocarbon group and the other with a butyl or pentyl group, it has been shown that a butyl group can be distinguished accurately from a pentyl group by a cyclopentyl group in aqueous ethanol.
Reductive Conversion of Chlorodifluoromethylbenzene with Samarium(II) Diiodide
Authors: Yoshida Masato , Suzuki Daiki , Iyoda Masahiko
Published: April 2006

PhCF2Cl suffered readily one-electron reduction with SmI2 to give PhCF2 radical and chloride ion. The resulting PhCF2 radical was further reduced with SmI2 to PhCF2SmI2 which reacted with electrophile such as Me3SiCl. The initially formed PhCF2 radical could be trapped by olefin or acetylene before it was reduced to PhCF2SmI2.
New Type of Cyclodextrin Sandwiched Porphyrin
Authors: Kuroda Yasuhisa , Egawa Yuichiro , Seshimo Hidenori , Ogoshi Hisanobu
Published: April 2006

A porphyrin having two molecules of 2,3,6-permethyl-β-cyclodextrin was synthesized. The starting materials employed were the αβαβ atoropisomer or meso-tetrakis(o-aminophenyl)porphyrin and the diacidchloride of A,D-dicarboxyl derivative of 2,3,6-permethyl-β-cyclodextrin. As expected the product contained two isomers which were isolated by HPLC.
Stereoselective Photoinduced Electron Transfer of Zinc Myoglobin with Optically Active Viologens
Authors: Tsukahara Keiichi , Kimura Chieko , Kaneko Junko , Hara Tomoko
Published: April 2006

The excited triplet state of zinc myoglobin was preferentially quenched by (S,S)-isomers of optically active viologens, 1,1′-bis(1-phenylethylcarbamoylmethyl)-4,4′-bipyridimum (OAV2+) and 1,1′-bis(1-(1-naphthyl)ethylcarbamoylmethyl)-4,4′-bipyridinium (NOAV2+) ions: kq((S,S)-)/kq((R,R)-) = 1.5 for NOAV2+ and 1.3 for OAV2+ at 25 °C and an ionic strength of 0.02 mol dm−3.
Children's representations of the earth: A methodological comparison
Authors: Panagiotaki Georgia , Nobes Gavin , Banerjee Robin
Published: April 2006

Investigation of children's understanding of the earth can reveal much about the origins and development of scientific knowledge. Vosniadou and Brewer (1992) claim that children construct coherent, theory‐like mental models of the earth. However, more recent research has indicated that children's knowledge of the earth is fragmented and incoherent. By testing the influence of question type (open vs. forced‐choice questions) and medium (drawings vs. 3‐D models) on the responses of 6‐year‐olds (N=59), this study investigated whether, and how, methodological differences account for the discrepant findings of previous research. Both the use of drawings and of open questions (Vosniadou and Brewer's methods) were found to increase the apparent incidence of naïve mental models. Moreover, the combination of 3‐D models and forced‐choice questions elicited more scientifically correct responses and higher proportions of scientific and inconsistent mental models than the combination of drawings and open questions. It is argued that children know more about the earth than the mental model theorists claim, and that naïve mental models of the earth are largely artifactual.
Standardized Bending and Breaking Test for the Normal and Osteoporotic Metaphyseal Tibias of the Rat: Effect of Estradiol, Testosterone, and Raloxifene
Authors: Stürmer Ewa K , Seidlová-Wuttke Dana , Sehmisch Stephan , Rack Thomas , Wille Joern , Frosch Karl H , Wuttke Wolfgang , Stürmer Klaus M
Published: April 2006

The fracture of bone plays a key role in osteoporosis. BMD measurement, however, is only an indirect parameter of this phenomenon. We therefore developed a highly sensitive three-point bending test for the metaphyseal tibias in rats to evaluate stiffness and strength. This was validated in a right-left comparison and a bioassay with soy-free food, estradiol, raloxifene, and testosterone in orchidectomized rats.

Introduction: Osteoporosis becomes manifest predominantly in the metaphyseal rat tibia. The anti-osteoporotic character of substances should, therefore, be tested (mechanically) in this bone area.

Materials and Methods: We evaluated a new three-point bending test for the metaphyseal tibia in rats in a right-left trial. In an animal experiment, we studied the change of bone quality under estradiol (E)-, raloxifene (R)-, and testosterone (T)-supplemented food and compared it with trabecular BMD (qCT).

Results: In the right-left comparison, the mean difference between the metaphyseal loads of both tibias in 37 rats was 8.43% for the maximum load (Fmax) and 6.46% for the failure load (fL). These results show the high reproducibility of the test, because they are close to the usual intraindividual difference of the two extremities. In a second experiment, four groups of 11 3-month-old male orchidectomized rats were fed with soy-free food only (C) or with the additives E, T, or R for 12 weeks. E and R were similar for Fmax and fL. There were significant differences in the stiffness (E = 406.92 N/mm versus R = 332.08 N/mm), the yield load (yL; E = 99.17 N versus R = 83.33 N), and the ratio between yL and Fmax (E = 86.33% versus R = 76.37%). T was similar to the controls concerning Fmax, fL, and stiffness. There were significant differences in yL (T = 49.00N versus C = 39.5N) and the ratio between yL and Fmax (T = 64.28% versus C = 51.28%).

Conclusions: Estradiol is superior to raloxifene concerning stiffness and yield load, and both are superior to testosterone. We conclude that the described three-point bending test for the metaphyseal tibia is a highly sensitive method to study hormones and substances with regard to their osteoprotective character. The precision and the low SD of the presented results are superior to the data from qCT and the calculated index of stiffness (SSI).
Immunocompetent Properties of Human Osteoblasts: Interactions With T Lymphocytes
Authors: Stanley Katherine T , VanDort Christa , Motyl Christopher , Endres Judith , Fox David A
Published: April 2006

We sought to determine whether osteoblasts (OBs) can serve as accessory cells (ACs) for T-cell activation and whether T cells directly activate OB production of IL-6, using primary human OBs (NHOst), the transformed fetal osteoblast line hFOB1.19, and an osteosarcoma line SaOS-2. Robust, bidirectional activating interactions were shown using each of these three human ostoblast lines.

Introduction: Osteoblasts (OBs) could come into contact with lymphocytes during inflammatory joint destruction and fracture repair.

Materials and Methods: We used several in vitro assays to assess the ability of T cells and OBs to interact in the generation of immune and inflammatory responses.

Results: By flow cytometry, three OB cell lines all were found to express ligands for T-cell co-stimulation. The integrin ligand CD54/ICAM-1 was constitutively expressed by hFOB1.19 and NHOst and was upregulated on SaOS-2 by IFN-γ. MHC Class II was upregulated on all three lines by IFN-γ. CD166/ALCAM, a ligand of the T-cell molecule CD6, was constitutively expressed on all three lines. A second putative CD6 ligand designated 3A11 was expressed on hFOB1.19 and NHOst, but not consistently on SaOS-2. The ectoenzyme CD26 (dipeptidyl peptidase IV) was expressed on hFOB1.19 and NHOst, but not on SaOS-2. All three cell lines presented superantigen to T cells, especially after treatment with IFN-γ. Superantigen presentation was inhibited by antibodies to the leukocyte integrin CD11a/CD18 (LFA-1), MHC Class II, and CD54/ICAM-1. T cells, particularly when cytokine activated for 7 days before co-culture, stimulated all three osteoblast lines to produce interleukin (IL)-6, and this effect was boosted when IL-17 was added to the co-cultures with either resting T cells or cytokine-activated T cells.

Conclusions: Bidirectional activating interactions are readily shown between human T cells and several types of human OBs. The expression by OBs of ligands for the T cell-specific molecule CD6, as well as other molecules involved in immune interactions, strongly suggests that such in vitro interactions are representative of physiologic or pathologic events that occur in vivo.
Increased Muscle Mass With Myostatin Deficiency Improves Gains in Bone Strength With Exercise
Authors: Hamrick Mark W , Samaddar Ted , Pennington Catherine , McCormick John
Published: April 2006

We tested the hypothesis that increased muscle mass augments increases in bone strength normally observed with exercise. Myostatin-deficient mice, which show increased muscle mass, were exercised along with wildtype mice. Results indicate that increases in bone strength with exercise are greater in myostatin-deficient mice than in wildtype mice, suggesting that the combination of increased muscle mass and physical activity has a greater effect on bone strength than either increased muscle mass or intense exercise alone.

Introduction: Muscle (lean) mass is known to be a significant predictor of peak BMD in young people, and exercise is also found to increase bone mass in growing humans and laboratory animals. We sought to determine if increased muscle mass resulting from myostatin deficiency would enhance gains in bone strength that usually accompany exercise.

Materials and Methods: Male mice lacking myostatin (GDF-8) were used as an animal model showing increased muscle mass. Wildtype and myostatin-deficient mice (n = 10-12 per genotype) were exercised on a treadmill for 30 minutes/day, 5 days/week, for 4 weeks starting at 12 weeks of age. Caged wildtype and myostatin-deficient mice (n = 10-12 per genotype) were included as sedentary controls. Structural and biomechanical parameters were measured from the radius.

Results: Ultimate force (Fu), displacement (Du), toughness (energy-to-fracture; U), and ultimate strain (εu) increased significantly with exercise in myostatin-deficient mice but not in normal mice. When Fu is normalized by body mass, exercised myostatin-deficient mice show an increase in relative bone strength of 30% compared with caged controls, whereas exercised wildtype mice do not show a significant increase in ultimate force relative to caged controls. Relative to body weight, the radii of exercised myostatin-deficient mice are >25% stronger than those of exercised normal mice.

Conclusions: Increased muscle mass resulting from inhibition of myostatin function improves the positive effects of exercise on bone strength.
Biochemical Markers of Bone Resorption Compared with Estimates of Bone Resorption from Radiotracer Kinetic Studies in Osteoporosis
Authors: Dr. Eastell R. , Colwell A. , Hampton L. , Reeve J.
Published: April 2006

The pyridinium cross-links of collagen pyridinoline (Pyd) and deoxypyridinoline (Dpd) are released during bone resorption and are neither metabolized nor absorbed from the diet. The aim of this study was to validate their use in osteoporosis. We studied 19 women with osteoporosis and estimated the bone resorption rate from a combined calcium balance/kinetics technique without (R) and with partial (RH) and “complete” (Res) correction for long-term exchange. The strongest correlation was observed between the bone-specific marker (Dpd) and with complete correction for long-term exchange (Res) (r = 0.71, p < 0.001). The intercept was not different from zero, suggesting that bone was the major source for Dpd. The crude ratio of Dpd to Res in the 19 women was 54.5; but the regression coefficient relating Dpd as the dependent variable to Res was 31.8 (95% CI 15.6–48.0), which was higher, but not significantly, than the ratio between Dpd and calcium (16.4) in 10 bone samples (cortical and trabecular bone). The weakest correlations between a biochemical marker and a kinetic index were those between hydroxyproline (a nonspecific marker of bone resorption) and R or RH. Treatment with hormone replacement therapy (HRT) or HRT and parathyroid hormone peptide 1–38 in seven women over 1 year resulted in similar percent changes in the biochemical markers and estimates of bone resorption. We conclude that the measurement of Dpd provides a reasonably accurate assessment of bone resorption in osteoporosis, and in the context of several repeat 24-h collections of urine offers measurement precision that is similar to that obtainable with methods depending on the use of radioisotopic tracers and the assessment of metabolic calcium balance.
Enhanced Nonsaturable Calcium Transport in the Jejunum of Rats During Lactation, but Not During Pregnancy
Authors: Boass Agna , Toverud Svein U.
Published: April 2006

The lactating (L) rat loses in excess of 100 mg of calcium (Ca) per day to milk at peak lactation. Most of the Ca must be provided by increased intestinal absorption. In an effort to examine adaptation of intestinal calcium absorption during lactation, nonsaturable absorption from the small intestine of rats was calculated from the disappearance of Ca from in situ ligated loops of jejunum during the last week of pregnancy and throughout lactation and weaning. Efficiency of absorption is reflected by the slope of the regression line of Ca absorbed on Ca introduced into the loop. Absorption of Ca in the jejunum was markedly enhanced starting at 5 days of lactation and for the remainder of lactation. Two days after weaning, the efficiency of jejunal Ca absorption decreased to the nonmated (NM) control level, while the lactation-associated intestinal hypertrophy persisted beyond 2 days postweaning. The percentages of water and Ca absorbed were positively and significantly correlated in both L and NM rats. In contrast to Ca, magnesium (Mg) and strontium (Sr) transport from ligated loops were not enhanced during lactation. Fifty millimolar glucose in the test solution increased the absorption of both water and Ca, but not Mg, from jejunal loops of NM rats. Glucose increased Ca absorption in NM rats up to the level seen in L rats. Glucose did not increase Ca absorption further in L rats, perhaps because of the greater availability of glucose to the intestine during lactation. We conclude that in rats the efficiency of nonsaturable Ca absorption from the jejunum is significantly increased during well established lactation, but not during pregnancy. The underlying mechanism appears to be specific for Ca, may be dependent on glucose, and is unrelated to intestinal hypertrophy.
Growth Hormone Normalizes Renal 1,25-Dihydroxyvitamin D3-24-Hydroxylase Gene Expression but Not Na+-Phosphate Cotransporter (Npt2) mRNA in Phosphate-Deprived Hyp Mice
Authors: Roy Stéphane , Martel Josée , Tenenhouse Harriet S.
Published: April 2006

The murine X-linked Hyp mutation is characterized by decreased renal expression of type II Na+-phosphate (Pi) cotransporter (Npt2) mRNA and an abnormal vitamin D response to Pi deprivation. The latter is manifest by an aberrant fall in serum 1,25-dihydroxyvitamin D3 (1,25(OH)2D) levels that is associated with an increase in renal 1,25(OH)2D-24-hydroxylase (24-hydroxylase), the first enzyme in the C-24 oxidation pathway. Because growth hormone (GH) enhances renal Na+-Pi cotransport and permits the adaptive 1,25(OH)2D response in Pi-deprived hypophysectomized rats, we examined the effects of GH on vitamin D metabolism and renal Npt2 mRNA abundance in Hyp mice fed control and low Pi diets. GH significantly decreased renal 24-hydroxylase activity (0.202 ± 0.020 to 0.098 ± 0.008 pmol/mg of protein/minute, p < 0.05) and mRNA abundance, relative to β-actin mRNA (299 ± 13 to 78 ± 14, p < 0.05), in Hyp mice fed the low Pi diet but had no effect on either parameter in mutants fed the control diet. Moreover, after GH treatment, renal 24-hydroxylase gene expression was no longer elevated in Pi-deprived Hyp mice relative to mutants fed control diet. In contrast, GH did not correct the serum concentration of 1,25(OH)2D in Pi-deprived Hyp mice. We also demonstrate that GH did not normalize renal Npt2 mRNA expression, relative to β-actin mRNA, in Hyp mice fed either control or low Pi diets. The present data demonstrate that normalization of renal 24-hydroxylase gene expression in Pi-deprived Hyp mice by GH is not sufficient to correct the serum concentration of 1,25(OH)2D and is not associated with an alteration in renal Npt2 mRNA expression.
Long-Term Treatment of Indomethacin Reduces Vertebral Bone Mass and Strength in Ovariectomized Rats
Authors: Saino Hitoshi , Matsuyama Toshikatsu , Takada Junichi , Kaku Tohru , Ishii Seiichi
Published: April 2006

We investigated the effect of the long-term treatment of indomethacin, on lumbar spinal bone mineral density (LSBMD), cancellous bone mass, structure, and strength of vertebral body in old ovariectomized (OVX) rats. Ten-month-old female Wistar rats were divided into five groups: the sham operated + vehicle (sham+VEH) group, the OVX + vehicle (OVX+VEH) group, the OVX + indomethacin (IN) 1.5 mg/kg/week (OVX+IN1.5) group, the OVX + IN 6.0 mg/kg/week (OVX+IN6.0) group, and the OVX + IN 15.0 mg/kg/week (OVX+IN15.0) group. IN or vehicle were given by subcutaneous injection (sc) three times per week. The treatments were started at 1 week after operation and continued for 24 weeks. LSBMD(L2–L5) was measured at 0, 12, and 24 weeks after the beginning of treatment. At the end of the experimental period, the animals were sacrificed, and bone histomorphometrical and biomechanical analysis of lumbar vertebral body were done. LSBMD, trabecular bone volume (BV/TV), and trabecular thickness (Tb.Th) decreased significantly in a dose-related manner with IN. In the OVX+IN15.0 group, LSBMD decreased by 12.7%, BV/TV decreased by 65.5%, and Tb.Th decreased by 32.8%, compared with the OVX+VEH group. In addition, the maximum stress in a compressive mechanical test of L4 vertebral body in OVX groups was also decreased in a dose-related manner with IN, and this value in the OVX+IN15.0 group was 31.3% lower than in the OVX+VEH group. We conclude that long-term treatment with IN accentuated the OVX-related decrease in trabecular bone mass and the compressive strength of lumbar vertebrae.
Comparison of Bone Density Measurement Techniques: DXA and Archimedes' Principle
Authors: Keenan Michael J. , Hegsted Maren , Jones Karen L. , Delany James P. , Kime James C. , Melancon Louis E. , Tulley Richard T. , Hong Kelly D.
Published: April 2006

The standard method for determination of density (g/cm3) of bones from small animals has been the application of Archimedes' principle. A recent development has been software for the determination of “density” (g/cm2) of small animal bones with dual-energy X-ray absorptiometry (DXA). We compared Archimedes' principle and DXA (Hologic QDR-2000) in the measurement of the densities of whole and hollowed femurs of 5- to 6-month-old retired female breeder rats. In an attempt to ensure detectable treatment differences, rats were used from a low-vitamin D Holtzman and a supplemental-vitamin D Sprague-Dawley colony. Whole femur densities were higher for supplemental-vitamin D colony rats than for low vitamin D rats using both techniques (Archimedes' principle, p < 0.002; DXA, p < 0.005), and the densities from the two techniques were highly correlated (r = 0.82, p < 0.0001). Actual density values were higher for Archimedes' principle than for DXA. Other variables such as femur ash weight and calcium content were also highly correlated to densities with both techniques. Hollowed femur density values were higher than whole femur values with Archimedes' principle but lower with DXA. Colony effects for hollowed femur densities were diminished with Archimedes' principle (p < 0.03) and eliminated with DXA (p < 0.53). Investigation of whole bones is more biologically relevant, and both techniques were effective in detecting differences between whole femurs from low-vitamin D and supplemental-vitamin D colony rats.
Calcitonin Alters Bone Quality in Beagle Dogs
Authors: Pienkowski David , Doers Thomas M. , Monier-Faugere Marie-Claude , Geng Zhaopo , Camacho Nancy P. , Boskey Adele L. , Malluche Hartmut H.
Published: April 2006

Because of its antiresorptive properties, calcitonin is widely used to prevent and treat osteoporosis. A stimulatory effect of calcitonin on osteoblasts has also been reported; however, a recent histologic study points to a negative effect of calcitonin on mineralization of cancellous bone. The present experiment was performed to determine whether the observed histological signs of alterations in mineralization are also observed in cortical bone and whether this results in changes in mechanical properties, mineral densities, or mineral properties of canine bone. Sixteen female adult beagle dogs were randomly allocated to receive either human calcitonin at a dose of 0.25 mg/dog (50 IU, n = 8) or vehicle (mannitol, n = 8) every other day for 16 weeks. At the end of the study, the dogs were euthanized. Both tibiae, L1 and L5 vertebrae, and iliac crest bone samples were excised and defleshed. Torsional mechanical properties of tibial diaphyses and compressive strengths of vertebrae were measured. Bone mineral densities (BMD) of tibiae and vertebrae were measured by using dual-energy X-ray absorptiometry. Ultrastructural mineral characteristics of iliac crest bone were determined by gravimetry and Fourier transform infrared spectroscopy (FTIR). Bone histomorphometry was performed in the cortical envelope of the iliac crest. Tibiae from dogs treated with calcitonin withstood significantly less maximum torque until failure, required less torsional energy to reach the maximum torque, and had less torsional stiffness than the tibiae from dogs treated with vehicle (p < 0.05). Cancellous cores of vertebrae from calcitonin-treated dogs withstood less compressive mechanical loading than did vertebral cores from vehicle-treated animals (p < 0.05). Dogs treated with calcitonin had less BMD of both tibiae and vertebrae than vehicle-treated animals (p < 0.05). Bones from calcitonin-treated dogs had significantly less ash content, which correlated with the lower phosphate-to-amide I (detected by FTIR) and greater carbonate-to-phosphate ratios than did bones from vehicle-treated dogs (p < 0.05). Calcitonin-treated dogs exhibited a decrease in bone formation and mineralization rates and an increase in mineralization lag time. These results point to a negative effect of calcitonin on bone quality. These findings are intriguing and call for further studies addressing whether the observed abnormalities are transient or permanent.
Reflection Ultrasound Velocities and Histomorphometric and Connectivity Analyses: Correlations and Effect of Slow-Release Sodium Fluoride
Authors: Zerwekh Joseph E. , Antich Peter P. , Mehta Shreefal , Sakhaee Khashayar , Gottschalk Frank , Pak Charles Y. C.
Published: April 2006

To better understand how structural and functional bone properties contribute to the changes in bone biomechanical properties revealed by ultrasound critical angle reflectometry (UCR) analysis, we measured both UCR velocities and histomorphometric properties in bone biopsy specimens from 33 osteoporotic patients before and following intermittent slow-release sodium fluoride (SRNaF) and continuous calcium citrate administration. Mean skeletal fluoride exposure was 17 months, and mean skeletal fluoride content was 0.203 ± 0.088 SD% bone ash. Intermittent SRNaF and continuous calcium citrate promoted significant increases in trabecular thickness (122 ± 18 SDμm to 131 ± 20, p = 0.020), mineral apposition rate (0.79 ± 0.26 to 1.05 ± 0.40 μm/day, p = 0.014), and a significant decline in eroded surface (3.9 ± 1.6 to 2.8 ± 1.4%, p = 0.002). There were also significant increases in node number (0.193 ± 0.100 to 0.368 ± 0.245, p < 0.01) and node-to-node strut length (0.076 ± 0.087 to 0.191 ± 0.173, p < 0.01) relative to total cancellous area. Cortical UCR velocity did not change but cancellous velocity significantly increased by 97 m/s following therapy (p = 0.0005). When compared against the significant changes in bone histomorphometry and connectivity, the sum of both cancellous and cortical ultrasound velocities was significantly correlated with node number/area (R2 = 0.305, p < 0.0001) and node-to-node strut length/area (R2 = 0.372, p < 0.0001) and to a lesser extent with mineral apposition rate (R2 = 0.106, p = 0.032). Multiple regression analysis demonstrated that 40% of the variance in the sum of the UCR velocities can be accounted for by the variability in these histomorphometric and connectivity parameters. There were no significant correlations between the sum of cortical and cancellous ultrasound velocities and cancellous bone volume (R2 = 0.014, p = 0.533), trabecular thickness (R2 = 0.012, p = 0.47), or bone mineral density (R2 = 0.003, p = 0.80). These observations indicate that velocity measurements with the UCR methodology show an improvement in bone elasticity associated, in part, with an improvement in the rate of bone mineralization and an improvement in bone quality at the structural level as shown by microarchitecture.
Temporal Changes of mRNA Expression of Matrix Proteins and Parathyroid Hormone and Parathyroid Hormone–Related Protein (PTH/PTHrP) Receptor in Bone Development
Authors: Kondo Hisatomo , Ohyama Takashi , Ohya Keiichi , Kasugai Shohei
Published: April 2006

Expression of parathyroid hormone and parathyroid hormone–related protein (PTH/PTHrP) receptor is one of the osteoblastic phenotypes; however, it has not been clear whether this phenotype expression is a marker of immature or mature osteoblasts. We examined the temporal expression pattern of PTH/PTHrP receptor in bone development in vivo and in vitro compared with the expression of other osteoblastic phenotypes: osteopontin (OPN), bone sialoprotein (BSP) and osteocalcin (OC), alkaline phosphatase (ALP), and mineralization. Total RNA was extracted from rat calvariae, and cell culture of rat bone marrow at different developmental stages and then Northern blot hybridization were performed. Mineralization was detected with contact microradiography (CMR) in calvaria or with Alizarin Red S staining in bone marrow cell culture. Both in calvaria and in marrow cell culture, extensive expression of OPN, BSP, type I collagen (COL I), and ALP coincided with the onset of mineralization, and OC expression was observed after mineralized tissue formation. Notably, PTH/PTHrP receptor was expressed at an early developmental stage (prenatal day 14 in calvaria, day 5 in culture) when mineralized tissue was not formed and other osteoblastic phenotypes were scarcely detected. Further study in cell culture revealed that the fold increase in cyclic adenosine monophosphate (cAMP) in response to PTH was elevated with the advance in the culture stage. These results indicate that mRNA expression of PTH/PTHrP receptor could be the early differentiation marker in osteoblastic lineage and that the levels of cAMP production in response to PTH represent the stage of osteoblastic differentiation.
Rapid Protein Kinase A—Mediated Activation of Cyclic AMP-Phosphodiesterase by Parathyroid Hormone in UMR-106 Osteoblast-like Cells
Authors: Ahlström Mikael , Dr. Lamberg-Allardt Christel
Published: April 2006

Parathyroid hormone (PTH) plays an essential role in osteoblast proliferation and differentiation. The effects of PTH are known to be mediated by cyclic adenosine monophosphate (cAMP) and calcium and by the activation of protein kinase C (PKC). cAMP is hydrolyzed to the inactive form 5′ AMP by cyclic nucleotide phosphodiesterases (PDEs). We have investigated the role of PTH on PDE regulation in UMR-106 osteoblast-like cells. Treatment with 10 nM PTH caused a 3-fold increase in the PDE activity. The activation of PDE could be seen within 2 minutes and reached maximal levels after 20 minutes. The PTH effect was dose dependent with a half-maximal dose of 2 nM. The effect of PTH could be mimicked by the cAMP analogs Bt2 cAMP and forskolin, but not by PTH fragment 3–34, calcium ionophore A23187, or by the PKC activator phorbol 12-myristate 13-acetate. The PDE activity stimulated by PTH could be abolished by the PKA inhibitor H-8. The PDE activated by PTH was inhibitable by low concentrations of the cAMP-PDE–specific inhibitor RO 20-1724 (IC50 = 0.2 μM), but not by low concentrations of the inhibitors of cGMP-stimulated and cGMP-inhibited PDEs MEP-1 and milrinone (IC50 for both compounds >30 μM). The PTH-stimulated cAMP accumulation was potentiated about 7-fold in the presence of RO 20-1724. H-8 potentiated the PTH-stimulated cAMP accumulation about 4-fold. Our results show that PTH rapidly stimulates the activity of cAMP-PDE in UMR-106 cells. The PDE activation involves cAMP and PKA. Inhibition of PKA can abolish the PTH-stimulated PDE activation and leads to increased accumulation of intracellular cAMP.
Establishment and Characterization of Conditionally Immortalized Stromal Cell Lines from a Temperature-Sensitive T-Ag Transgenic Mouse
Authors: Feuerbach Dominik , Loetscher Erika , Buerki Kurt , Sampath T. K. , Dr. Feyen Jean H. M.
Published: April 2006

We established bone marrow stromal cell lines from a transgenic mouse that harbors a temperature-sensitive mutant of the simian virus 40–derived large T-antigen under the control of an major histocompatibility complex (MHC) I promotor. These cell lines were screened for their ability to induce the formation of osteoclasts in a spleen cell/stromal cell coculture system. By means of this screen, five clones, referred to as marine bone marrow stromal clone 1 (mBMS-B1) mBMS-B2, mBMS-B14, mBMS-B18, and mBMS-B21, were selected for detailed characterization. Cell growth depends on culture conditions, i.e., cells grow at 33°C in the presence of murine interferon-γ, whereas cell proliferation ceases at 39°C. The phenotype of the cells is also correlated with the culture conditions because the osteoclast inductive capacity is only seen at 39°C, indicating that the cells undergo differentiation when the transforming agent is inactivated. These conditionally immortalized stromal cells can be induced to express a variety of markers that are typical for mature osteoblasts, e.g., alkaline phosphatase activity and expression of functional parathyroid hormone receptor after stimulation with soluble osteogenic protein 1 (sOP-1). mRNA analysis revealed the expression and regulation of osteopontin, osteonectin, and collagen α1(I) as well as the inducibility of osteocalcin upon treatment with sOP-1. The cells have the potential to form mineralized nodules in supplemented medium. We observed expression of vascular cell adhesion molecule-1, which is stimulated upon treatment of the cells with 1α,25-dihydrocholecalciferol after 4 days, indicating the presence of the receptor for this steroid. These cell lines represent a model to study mechanisms and factors involved in osteoblast differentiation.
Osteoblastic Proliferation in Bone Biopsies from Patients with End-Stage Chronic Renal Failure
Authors: Dr. Serrano Sergio , Mariñoso Maria L. , Torres Armando , Lorenzo Victor , Keysers Ursula , Lloreta Jose , Nacher Montserrat , Garcia Carmen , Ballester Jorge , Diez Adolfo , Aubia Jaume
Published: April 2006

Osteoblasts have traditionally been considered to be terminally differentiated cells and therefore unable to divide. Data in recent years, however, indicate that cellular differentiation does not usually preclude preservation of proliferative ability and that most differentiated cells are able to divide under adequate stimuli. The aim of this study was to assess whether cubic osteoblasts undergo proliferation during the formation phase of the remodeling cycle under a stimulus that increased bone turnover. For that purpose, the osteoblastic proliferation index (OPI) was analyzed by DNA image cytometry in transiliac bone biopsies from 33 patients with chronic renal failure (23 men, 10 women; mean age 50.4 ± 15.1 years) who have been classified into low (n = 13), normal (n = 15), and high (n = 15) bone turnover according to activation frequency (Ac.f). OPI was significantly higher (p < 0.002) in the high bone turnover group (13.90 ± 4.72%) compared with the low (2.38 ± 4.13%) and normal turnover groups (2.84 ± 4.04%). There was a positive correlation between OPI and the following histomorphometric parameters: bone formation rate, surface referent (r = 0.76, p = 0.00001), activation frequency (r = 0.73, p = 0.00001), mineral apposition rate (r = 0.73, p = 0.00001), bone formation rate, volume referent (r = 0.71, p = 0.00001), and mineralizing surface (r = 0.62, p = 0.0001). This study shows that a rise in bone turnover is associated with a marked increase of bone-forming cell proliferation in patients with end-stage chronic renal failure. From this finding, it may be concluded that cubic osteoblasts do not behave as “terminally differentiated” cells in vivo, because a high proportion of them are still able to divide.
Determinants of Radial Bone Density as Measured by PQCT in Pre- and Postmenopausal Women: The Role of Bone Size
Authors: Wapniarz Monika , Lehmann Reiner , Reincke Martin , Schönau Eberhard , Klein Klaus , Allolio Bruno
Published: April 2006

The aim of the present study was to examine the influence of anthropometric, hormonal, and geometric factors on the variability of radial bone mineral density (BMD) in women. In 583 healthy pre- and postmenopausal females (aged 40–60 years) radial total (BD) and trabecular BMD (TBD) was measured by peripheral quantitative computerized tomography at the nondominant distal forearm. In addition, 29 women who had suffered a Colles' fracture after minor trauma were also evaluated. There was no age-dependent change in radial BD and TBD before menopause. We found a negative correlation between BMD and age and years since menopause (YSM) in postmenopausal women (BD = 422.73 − 2.342 age − 6.308YSM; r = 0.36, p = 0.0001, n = 128). The variation of YSM, body mass index (BMI), and age accounted for 20% of the variability of BD in postmenopausal women. In contrast, in premenopausal women, only 3% of the variability could be explained by anthropometric variables. Bone mineral content (BMC) and bone area, but not BMD at the distal radius, were significantly correlated to grip strength (r = 0.25, p = 0.006 for BMC, r = 0.26, p = 0.003 for area). The cross-sectional bone area of the CT slice showed a significant increase with aging (y = 263.02 + 1.25x; r = 0.14, p = 0.0009). There was a strong negative correlation between bone area and BD and TBD (y = 516.04 − 0.668x; r = −0.57, p < 0.0001 for BD). If BMD is normalized for BA, variation is reduced by 32% (for BD) and 10% (for TBD), respectively. Women with Colles' fracture had a significantly lower TBD normalized for BA (fracture group [−0.71 ± 0.88] vs. normals [0.03 ± 0.99]; p = 0.00009). Our results show that YSM and BMI are predictors of postmenopausal BMD. However, radial BMD is influenced strongly by geometric variables such as cross-sectional bone area.
Prostaglandin E2 (PGE2) and Risedronate Was Superior to PGE2 Alone in Maintaining Newly Added Bone in the Cortical Bone Site After Withdrawal in Older Intact Rats
Authors: Ma Yan Fei , Lin Bai Yun , Jee Webster S. S. , Lin Chao Hua , Chen Yong Yong , Ke Hua Zhu , Li Xiao Jian
Published: April 2006

The objects of this study were (1) to determine the effects of risedronate (Ris) and prostaglandin E2 (PGE2) alone and in combination, on tibial diaphyses of older intact female rats; and (2) to observe the fate of any extra bone if formed after withdrawal of the treatment. Nine-month-old female Sprague-Dawley rats were treated with 6 mg of PGE2/kg/day, 1 or 5 μg of Ris/kg twice a week, or 6 mg of PGE2/kg/day plus 1 or 5 μg of Ris/kg twice a week for the first 60 days and followed by vehicle injections for another 60 days. Cross-sections of double fluorescent labeled, undecalcified tibial diaphyses proximal to the tibiofibular junction were processed for histomorphometry. We found that: (1) neither the 1 μg nor the 5 μg of Ris treatment in the 60-day on/60-day off group showed any histomorphometric differences from age-related controls; (2) while the 60 days of PGE2 treatment added extra cortical bone (6%) on the tibial shaft (due to stimulation of periosteal, endocortical, and marrow trabecular bone formation), the new endocortical and most of the new marrow trabecular bone were lost when treatment was withdrawn; however, the new periosteal bone remained; (3) PGE2 with Ris added the same amount of new bone to tibial diaphysis as did PGE2 alone and upon withdrawal, new marrow trabecular bone was lost but new periosteal and endocortical bones were preserved in PGE2 + 1 μg of Ris on/off group. In contrast, all the new bone was maintained in the PGE2 + 5 μg of Ris on/off group; (4) PGE2 + Ris cotreatment failed to block the increase in cortical bone porosity induced by PGE2; and (5) in the PGE2 alone and PGE2 + 1 μg of Ris on/off groups bone turnover was higher than that in the PGE2 + 5 μg of Ris on/off group. These results indicate that on/off treatment with PGE2 and Ris is superior to PGE2 alone in that it forms the same amount of new bone during treatment, but preserves more cortical bone during withdrawal. Depression of bone resorption and turnover were the tissue mechanisms responsible for this protection.
Phenotypic Heterogeneity of Osteoblast-like MC3T3-E1 Cells: Changes of Bradykinin-Induced Prostaglandin E2 Production During Osteoblast Maturation
Authors: Dr. Leis Hans J. , Hulla Wolfgang , Gruber Reinhard , Huber Evelyn , Zach Doris , Gleispach Helmut , Windischhofer Werner
Published: April 2006

We have examined clonal murine calvarial MC3T3-E1 cells obtained from different sources to compare their osteoblastic features (alkaline phosphatase [ALP], cyclic adenosine monophosphate [cAMP] response to parathyroid hormone, prostaglandin E2 (PGE2) and PGE1, bradykinin-induced production of PGE2). It was found that the sublines investigated showed large variation of the above-mentioned parameters, which may be attributed to distinct differentiated stages of osteoblast development. Increase of ALP activity was paralleled by an increase in cAMP accumulation in response to the above-mentioned agents. The most striking difference was observed with bradykinin-induced production of PGE2. Early stage cells (low ALP) produced high levels of PGE2, whereas cells with high ALP activity showed no bradykinin stimulation at all. This was consistent with the results of specific binding of3H-bradykinin to its receptor and also correlated well with the bradykinin-induced signal transduction sequence (inositol triphosphate liberation and elevation of intracellular calcium levels). This was confirmed by Northern blot analysis of bradykinin receptor mRNA expression. These results indicate that the widely used osteoblast-like cell line MC3T3-E1 is synonymous for multiple sublines, representing different stages of osteoblast development. These sublines were most likely emerging from the early stage cell line due to the applied culture conditions. Moreover, distinct biochemical features are displayed in correlation to the differentiation stage, thus providing a useful model to study the molecular mechanism of osteoblast maturation.