Effects of CO2 laser irradiation of the gingiva during tooth movement.

Patients often feel pain or discomfort in response to orthodontic force. It was hypothesized that CO(2) laser irradiation may reduce the early responses to nociceptive stimuli during tooth movement. The distribution of Fos-immunoreactive (Fos-IR) neurons in the medullary dorsal horn of rats was evaluated. Two hrs after tooth movement, Fos-IR neurons in the ipsilateral part of the medullary dorsal horn increased significantly. CO(2) laser irradiation to the gingiva just after tooth movement caused a significant decrease of Fos-IR neurons. PGP 9.5- and CGRP-positive nerve fibers were observed in the PDL of all study groups. The maximum temperature below the mucosa during CO(2) laser irradiation was less than 40 degrees C. It was suggested that CO(2) laser irradiation reduced the early responses to nociceptive stimuli during tooth movement and might not have adverse effects on periodontal tissue.

Construction of Lactobacillus plantarum strain with enhanced L-lysine yield.

AIM: To enhance L-lysine secretion in Lactobacillus plantarum. METHODS AND RESULTS: An S-2-aminoethyl-L-cystein (AEC)-resistant mutant of L. plantarum was isolated, and it produced L-lysine at considerably higher level than the parent strain. Aspartokinase in the mutant has been desensitized to feedback inhibition by L-lysine. The nucleotide sequence analysis of thrA2 that codes for aspartokinase in the mutant predicted a substitution of glutamine to histidine at position 421. L-Lysine-insensitive aspartokinase, together with aspartate semialdehyde dehydrogenase, dihydrodipicolinate synthase, and dihydrodipicolinate reductase genes, was cloned from L. plantarum DNA to a shuttle vector, pRN14, and the genes were then transformed individually into the AEC-resistant mutant and the parent strain. The overexpression of the genes led to the increase in the activity of enzymes they encode in vitro. However, only the strain overexpressing aspartokinase or dihydrodipicolinate synthase produced more L-lysine. CONCLUSIONS: The desensitization of aspartokinase to L-lysine in L. plantarum led to the overproduction of L-lysine. The overexpression of L-lysine-insensitive aspartokinase or dihydrodipicolinate synthase enhanced L-lysine secretion in L. plantarum. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of the L-lysine-overproducing strain of L. plantarum in food or feed fermentation may increase the L-lysine content of fermented products.

Denervation resulting in dento-alveolar ankylosis associated with decreased Malassez epithelium.

Inferior alveolar nerve denervation causes appreciable decreases in the distribution of epithelial rests of Malassez. To explore roles of the Malassez epithelium, we attempted to evaluate possible changes in dento-alveolar tissues surrounding this epithelium by experimental denervation. We found that denervation led to dento-alveolar ankylosis with a decrease in the width of the periodontal spaces. Interestingly, with regeneration of the Malassez epithelium 10 weeks after the denervation, the periodontal space width showed a correspondingly significant increase. These findings suggest that the Malassez epithelium may be involved in the maintenance of periodontal space and that sensory innervation might be indirectly associated with it. In addition, it is of interest that denervation activated root resorption of the coronal root surface and that the consequently resorbed lacunae were repaired by cellular cementum. It is suggested that Malassez epithelium may negatively regulate root resorption and induce acellular cementum formation.

A developmentally regulated and psychostimulant-inducible novel rat gene mrt1 encoding PDZ-PX proteins isolated in the neocortex.

Single or repeated exposure to psychostimulants such as amphetamines and cocaine after postnatal week 3 leads to an enduring enhancement in the psychotomimetic responses elicited by a subsequent challenge of a stimulant in rodents. This behavioral sensitization phenomenon has been considered to be the neural consequences of stimulant-induced alterations in gene expression in the brain after a critical period of postnatal development. Using a differential cloning technique, RNA arbitrarily primed PCR, we have now identified from the rat neocortex a novel and developmentally regulated methamphetamine (MAP)-inducible gene mrt1 (MAP responsive transcript 1). mrt1 encodes two major types of PDZ- and PX-domains containing proteins of approximately 62 kDa in size with different carboxy termini, Mrt1a and Mrt1b. The mrt1 mRNAs for Mrt1a, mrt1a, and for Mrt1b, mrt1b, are predominantly expressed in various brain regions and the testes, respectively. Acute MAP injection upregulated mrt1b expression in the neocortex after postnatal week 3 in a D1 receptor antagonist-sensitive manner without affecting mrt1a expression. This upregulation was mimicked by another stimulant, cocaine, whereas pentobarbital and D1 antagonist failed to change the mrt1b transcript levels. Moreover, repeated daily treatment of MAP, but not MAP plus D1 antagonist, for 5 days caused an augmentation of the basal expression of mrt1b 2 and 3 weeks after the drug discontinuation. These late-developing, cocaine-crossreactive, D1 antagonist-sensitive and long-term regulations of mrt1b by MAP are similar to the pharmacological profiles of stimulant-induced behavioral sensitization, and therefore may be associated with the initiation and/or maintenance of the long-term neuronal adaptation.

In vivo conversion of a glycan to human compatible type by transformed tobacco cells.

Horseradish peroxidase isozyme C (HRP; EC 1.11.1.7) was used as a model protein to evaluate the capacity of tobacco cells transformed with human beta 1,4-galactosyltransferase (GT6) to modify and galactosylate a foreign glycoprotein. Cells transformed with the HRP gene are designated as BY2-HRP and GT6-HRP, for wild type BY2 and GT6 transformed cells, respectively. Expression of HRP cells was confirmed by isoelectric focusing, peroxidase activity staining, Western blotting, and enzymatic assays. The presence of HRP galactosylated N-glycans in GT6-HRP cells was analyzed by lectin staining, affinity chromatography, and structural analyses of pyridylamino-labeled RCA(120)-bound sugar chains. The structure of Gal(1)GlcNAc(1)Man(5)GlcNAc(2) was proposed based from the results of exoglycosidase digestions and two-dimensional sugar chain mapping. Unlike the HRP produced in BY2-HRP cells, the HRP from GT6-HRP cells has galactosylated glycoproteins that did not bind to the xylose-specific antiserum, suggesting the absence of the beta 1,2-xylose residue in the sugar chain.

Production and characterization of active soluble human beta1,4-galactosyltransferase in Escherichia coli as a useful catalyst in synthesis of the Gal beta1-->4 GlcNAc linkage.

An active and soluble human beta1,4-galactosyltransferase (beta-GT) was produced in Escherichia coli using a maltose-binding protein fusion system. The purified recombinant beta-GT has a K(m) value of 0.035 mM for UDP-galactose and a V(max) of 643 x 10(3) nmol/mg/h. The enzyme catalyzes the transfer of galactose from UDP-galactose to N-linked oligosaccharides. The properties of the purified enzyme were identical to those of bovine milk beta-GT.

Effect of alpha1,2-mannosidic linkage located in a alpha1,3-branch of Man6GlcNAc2 oligosaccharide on enzyme activity of recombinant human Man9-mannosidase produced in Escherichia coli.

We produced the human kidney Man9-mannosidase in Escherichia coli and studied the effect of the alpha-1,2-mannosidic linkage located in the alpha branch of Manalpha1-6(Manalpha1-2Manalpha1-3)Manalpha1-6(Manalpha1-3)Manbeta1-4GlcNAcbeta1-4-GlcNAc, an N-linked oligosaccharide, on the enzyme activity. The alpha1,2-mannose residue influenced the rate of hydrolysis of and substrate preference for other alpha-1,2-mannosidic linkages.

Identification of putative gene encoded on ORF16 of the 81 kb contig of Arabidopsis thaliana chromosome III as alpha-mannosidase.

We isolated cDNA corresponding to open reading frame (ORF) 16 of the 81 kb contig of Arabidopsis thaliana chromosome III [Quigley., Nucleic Acids Res., 24, 4313-4318 (1996)] and expressed alpha-mannosidase activity in tobacco suspension-cultured cells, which revealed that ORF16 encodes alpha-mannosidase. We also suggested that Arabidopsis harbors three genes encoding alpha-mannosidase by homology search against the database.

Human N-acetylglucosaminyltransferase I. Expression in Escherichia coli as a soluble enzyme, and application as an immobilized enzyme for the chemoenzymatic synthesis of N-linked oligosaccharides.

N-Acetylglucosaminyltransferase I (GnT-I), which catalyzes the transfer of an N-acetylglucosamine residue from UDP-N-acetylglucosamine to the alpha1,3-linked mannose on Man5GlcNAc2 (M5), is a critical enzyme for the synthesis of high-mannose-type to complex-type glycan structures in N-linked glycan processing. We developed a large-scale preparation system for recombinant human GnT-I (hGnT-I) using the maltose binding protein (MBP) fusion system to facilitate the chemoenzymatic route for complex-type N-linked glycan synthesis. MBP-fused GnT-I was purified by affinity chromatography on an amylose resin column. The relative activity of MBP-fused GnT-I toward high-mannose-type N-linked oligosaccharides was 100% for Man5GlcNAc2, 52% for Man3GlcNAc2, 17% for Man6GlcNAc2. MBP-fused GnT-I exhibited optimal activity at pH 6.5-9.5 and was more active between pH 6.5-9.0. The optimum temperature for MBP-fused GnT-I activity was 40 degrees C, but the enzyme was active between 0-70 degrees C. Mn2+ and Co2+ were critical for the enzyme activity, while Zn2+ and Ca2+ inhibited the activity. Kinetic analysis of the purified enzyme showed an apparent K(m) value of 0.483 mM and a V(max) of 101 nmol/mg/min for M5. Immobilization of MBP-fused GnT-I on the amylose resin led to an 80% yield of the high mannose-type-of oligosaccharide.

Glycoproteins secreted from suspension-cultured tobacco BY2 cells have distinct glycan structures from intracellular glycoproteins.

Glycan structures of glycoproteins secreted in the spent medium of tobacco BY2 suspension-cultured cells were analyzed. The N-glycans were liberated by hydrazinolysis and the resulting oligosaccharides were labeled with 2-aminopyridine. The pyridylaminated (PA) glycans were purified by reversed-phase and size-fractionation HPLC. The structures of the PA sugar chains were identified by a combination of the two-dimensional PA sugar chain mapping, MS analysis, and exoglycosidase digestion. The ratio (40:60) of the amount of glycans with high-mannose-type structure to that with plant-complex-type structure of extracellular glycoproteins is significantly different from that (ratio 10:90) previously found in intracellular glycoproteins [Palacpac et al., Biosci. Biotechnol. Biochem. 63 (1999) 35-39]. Extracellular glycoproteins have six distinct N-glycans (marked by *) from intracellular glycoproteins, and the high-mannose-type structures account for nearly 40% (Man5GlcNAc2, 28.8%; Man6GlcNAc2*, 6.4%; and Man7GlcNAc2*, 3.8%), while the plant-complex-type structures account for nearly 60% (GlcNAc2Man3Xyl1GlcNAc2*, 32.1%; GlcNAc1Man3Xyl1GlcNAc2 (containing two isomers)*, 6.2%; GlcNAc2Man3GlcNAc2*, 4.9%; Man3Xyl1Fuc1GlcNAc2, 8.3%; and Man3Xyl1GlcNAc2, 3.7%).

Etidronate inhibits human osteoblast apoptosis by inhibition of pro-apoptotic factor(s) produced by activated T cells.

Humoral factors produced by activated T cells are thought to be important in the development of bone loss in patients with rheumatoid arthritis (RA). We investigated the inhibitory effect of etidronate disodium (EHDP) on apoptosis of human osteoblasts induced by supernatants from in vitro activated T cell cultures. Human osteoblastic cell line MG63 cells and human primary osteoblast-like cells were used in the present study as human osteoblasts. T cells were incubated with interleukin-2 and further activated with 1 2-o-tetradecanoyl-phorbol 13-acetate and ionomycin, either in the presence or absence of EHDP. After we carried out the cultivation, we examined the cytotoxicity of cultured T cell supernatants toward MG63 cells and human primary osteoblast-like cells. Supernatants from activated but not resting T cell cultures efficiently induced apoptosis of MG63 cells and primary osteoblast-like cells. Supernatants from activated T cell cultures, incubated with EHDP, exhibited significantly less cytotoxicity than did supernatants incubated in the absence of EHDP. In contrast, the cytotoxicity of activated T cell culture supernatants was not affected by direct treatment of human osteoblasts with EHDP. The concentration of soluble Fas ligand in activated T cell culture supernatants was actually increased by EHDP. However, EHDP did not influence soluble Fas and tumor necrosis factor-alpha concentrations in the supernatant. Furthermore, treatment of human osteoblasts with EHDP did not alter their expression of Bcl-2/Bcl-xL or their sensitivity to anti-Fas immunoglobulin M-induced apoptosis. Our results suggest that EHDP inhibits the production of soluble factor that induces apoptosis of human osteoblasts and thus exhibits a protective action toward human osteoblast apoptosis induced by activated T cell culture supernatants. Although the exact EHDP-regulated molecule that induces apoptosis of human osteoblasts is unknown at present, our study may explain part of the therapeutic action of bisphosphonates in RA complicated by bone loss.

High expression of the penicillin G acylase gene (pac) from Bacillus megaterium UN1 in its own pac minus mutant.

By marker exchange mutagenesis, Bacillus megaterium strain UN-1 (Bm-UN1) was used to prepare a mutant strain B. megaterium UN-cat (Bm-UNcat) lacking the penicillin G acylase gene (pac). The pac gene from Bm-UN1 was subcloned into pTF6 and the resultant plasmid, pBA402, was introduced into Bm-UNcat and Bacillus subtilis. Bm-UNcat harbouring pBA402 produced high penicillin G acylase (PAC) activity of 13.7, 19.5 and 20.4 U ml(-1) at 24, 36 and 48 h of culture, respectively. This was two- to fivefold higher than PAC produced by B. subtilis harbouring pBA402 and about 20-fold higher than PAC produced by the parent strain, Bm-UN1.

Inferior alveolar nerve transection inhibits increase in osteoclast appearance during experimental tooth movement.

To evaluate the role of sensory nerve innervation in alveolar bone remodeling during experimental tooth movement, we investigated histomorphometrically the influence of sensory nerve denervation on bone metabolism. Seven days after inferior alveolar nerve (IAN) transection or a sham operation in rats, orthodontic force was applied to the animals by inserting an elastic module interproximally between the lower first molar and second molar. Twenty-four hours after the application of the orthodontic force, osteoclast number, osteoclast surface, and osteoblast surface were measured on the trabecular bone surface in the interradicular septum of the lower second molar. The distribution of sensory nerve fibers immunoreactive to antibody against calcitonin gene-related peptide (CGRP) was also evaluated. In the sham-operated rats, CGRP-immunoreactive nerves were observed to be distributed along the blood vessels in the trabecular alveolar bone. Experimental tooth movement resulted in a fivefold increase in the number of osteoclasts and in increased immunoreactivity of nerves to anti-CGRP in the trabecular bone. However, IAN transection depleted the immunoreactivity to anti-CGRP and reduced the osteoclast number and osteoclast surface significantly. On the other hand, in the rats that were not subjected to experimental tooth movement, there was no significant difference in osteoclast number between sham-operated and IAN-transected rats. Significant changes were not observed in osteoblast surfaces associated with experimental tooth movement or nerve transection. These findings suggest that sensory nerves play an important role in regulating bone resorptive activity during experimental tooth movement.

Epithelial rests of Malassez express immunoreactivity of TrkA and its distribution is regulated by sensory nerve innervation.

The periodontal ligament is the connective tissue that fills the space between the tooth and its bony socket. It is abundantly innervated by the sensory and sympathetic nerves. We first investigated the immunoreactivity of TrkA, which is a high-affinity receptor of nerve growth factor (NGF), in the periodontal ligament of rats. Immunoreactivity was observed at the epithelial cells in the cervical and furcation regions of the molars. These epithelial cells, which gather together to form clusters or networks, are known as the epithelial rests of Malassez. Immunoreactivity was not observed in other non-neuronal cells, such as osteoblasts, fibroblasts, odontoblasts, cementoblasts, endothelial cells, and/or osteoclasts. On the basis of these findings, we investigated the possible involvement of sensory nerve innervation in the immunoreactivity of the epithelial cells. Denervation of the inferior alveolar nerve resulted in a marked decrease in the distribution area and size of the clusters of immunoreactive cells compared with those of sham-operated rats. These findings suggest that sensory nerve innervation may have a regulatory role in maintenance of the epithelial rests of Malassez expressing TrkA in the periodontal ligament.

Parathyroidectomy for primary hyperparathyroidism induces positive uncoupling and increases bone mineral density in cancellous bones.

OBJECTIVE: Osteopenia is an important feature of primary hyperparathyroidism (PHP). However, little is known about the change of bone mineral density (BMD) in PHP after surgery. The aim was to investigate the mechanisms of increased BMD after parathyroidectomy in patients with PHP. DESIGN: Prospective observational study. PATIENTS: Ten patients with PHP (7 women, 3 men; mean age 53.2+/-9.1 years). All patients underwent parathyroidectomy for excision of parathyroid adenoma. MEASUREMENTS: BMDs of two cancellous bone-rich sites (L2-L4 lumbar spine and ultra-distal end of the radius, RUD) and one cortical bone-rich site (distal third of the radius, R33%) were measured using dual energy X-ray absorptiometry, before, and 3, 6 and 12 months after surgery. Serum intact PTH, intact osteocalcin, bone type alkaline phosphatase (b-ALP), alkaline phosphatase, calcium, and urinary deoxypyridinoline (Dpd) were measured before, and 1 and 3 days, and 1, 2, 3, 4, 6, 8, 12, and 24 weeks after surgery. RESULTS: Parathyroidectomy resulted in a significant increase in BMDs of L2-L4 and RUD at 3 months postoperatively. Urinary Dpd levels decreased within a few days after surgery, while b-ALP and osteocalcin decreased more slowly throughout the first few months after surgery. The ratio of osteocalcin/Dpd at 1 week after surgery correlated significantly with the percentage change in BMD of L2-L4 at 3 and 6 months after surgery. The ratio of osteocalcin/Dpd at 2 weeks correlated significantly with the percentage change in BMD of L2-L4 at 3, 6 and 12 months after surgery. The preoperative values of osteocalcin, b-ALP, PTH and calcium were positively correlated with the change in BMD of RUD at 3 months and L2-L4 at 12 months, RUD at 6 months, RUD at 3 months and L2-L4 at 12 months, respectively. CONCLUSIONS: In primary hyperparathyroidism patients, the major increase in bone mineral density following parathyroidectomy occurs within 3 months. Parathyroidectomy resulted in a marked increase in bone mineral density of cancellous bones compared to that of cortical bones. The early increase in bone mineral density was due to a preferential activation of bone formation over bone resorption as evidenced by changes in bone metabolic markers. Our results also showed that the preoperative levels of bone metabolic markers may predict the gain in bone mineral density after parathyroidectomy.

Tumor necrosis factor-alpha and interleukin-1beta increase the Fas-mediated apoptosis of human osteoblasts.

Our recent work demonstrated functional Fas expression on human osteoblasts, and the histologic examination of the periarticular osteoporosis region in patients with rheumatoid arthritis (RA) showed apoptosis in osteoblasts. High concentrations of interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha), and IL-6--which are thought to increase bone resorption--have been determined in RA synovium. We investigated the effect of these cytokines on the Fas-mediated apoptosis of human osteoblasts. The human osteoblastic cell line MG63 and human primary osteoblast-like cells from bone biopsy specimens were used as human osteoblasts. Fas expression on these cells was examined by flow cytometry, and Fas-mediated apoptosis induced by anti-Fas immunoglobulin M (IgM) was determined by a chromium 51 release assay, the presence of cells with hypodiploid DNA, staining with Hoechst 33258 dye, and the detection of DNA fragmentation on agarose gel electrophoresis. The proliferation of osteoblasts was analyzed by a tritiated thymidine incorporation assay. Spontaneous apoptosis was not found on cultured osteoblasts. The apoptosis of human osteoblasts was not induced by TNF-alpha, IL-1beta, or IL-6 alone in the absence of anti-Fas IgM. In addition, proliferation of the cells was not affected by these cytokines. Fas was constitutively expressed on unstimulated osteoblasts, and treatment of these cells with IL-1beta or TNF-alpha significantly augmented Fas expression. Human osteoblasts were committed to apoptosis with anti-Fas IgM, and the treatment of both IL-1beta and TNF-alpha markedly increased Fas-mediated apoptosis. TNF-alpha augmented both Fas expression and Fas-mediated apoptosis more efficiently than did IL-1beta. In addition, an additive effect on both Fas expression and Fas-mediated apoptosis was demonstrated when TNF-alpha and IL-1beta were added to osteoblasts. IL-6 influenced neither Fas expression nor the Fas-mediated apoptosis of osteoblasts. Furthermore, no synergistic effect of IL-6 with IL-1beta or TNF-alpha was observed. IL-1beta, TNF-alpha, or IL-6 did not change Bcl-2 expression. Our results suggest that IL-1beta and TNF-alpha regulate osteoblast cell number by up-regulating the Fas-mediated apoptosis of osteoblasts, one of the putative mechanisms inducing periarticular osteoporosis in patients with RA.

Stable expression of human beta1,4-galactosyltransferase in plant cells modifies N-linked glycosylation patterns.

beta1,4-Galactosyltransferase (UDP galactose: beta-N-acetylglucosaminide: beta1,4-galactosyltransferase; EC 2.4.1. 22) catalyzes the transfer of galactose from UDP-Gal to N-acetylglucosamine in the penultimate stages of the terminal glycosylation of N-linked complex oligosaccharides in mammalian cells. Tobacco BY2 cells lack this Golgi enzyme. To determine to what extent the production of a mammalian glycosyltransferase can alter the glycosylation pathway of plant cells, tobacco BY2 suspension-cultured cells were stably transformed with the full-length human galactosyltransferase gene placed under the control of the cauliflower mosaic virus 35S promoter. The expression was confirmed by assaying enzymatic activity as well as by Southern and Western blotting. The transformant with the highest level of enzymatic activity has glycans with galactose residues at the terminal nonreducing ends, indicating the successful modification of the plant cell N-glycosylation pathway. Analysis of the oligosaccharide structures shows that the galactosylated N-glycans account for 47.3% of the total sugar chains. In addition, the absence of the dominant xylosidated- and fucosylated-type sugar chains confirms that the transformed cells can be used to produce glycoproteins without the highly immunogenic glycans typically found in plants. These results demonstrate the synthesis in plants of N-linked glycans with modified and defined sugar chain structures similar to mammalian glycoproteins.

Structures of N-linked oligosaccharides of glycoproteins from tobacco BY2 suspension cultured cells.

The structures of N-linked sugar chains of glycoproteins expressed in tobacco BY2 cultured cells are reported. Five pyridylaminated (PA-) N-linked sugar chains were derived and purified from hydrazinolysates of the glycoproteins by reversed-phase HPLC and size-fractionation HPLC. The structures of the PA-sugar chains purified were identified by two-dimensional PA-sugar chain mapping, ion-spray MS/MS analysis, and exoglycosidase digestions. The five structures fell into two categories; the major class (92.5% as molar ratio) was a xylose containing-type (Man3Fuc1 Xyl1GlcNAc2 (41.0%), GlcNAc2Man3Fuc1Xyl1GlcNAc2 (26.5%), GlcNAc1Man3Fuc1Xyl1GlcNAc2 (21.7%), Man3 Xyl1GlcNAc2 (3.3%)), and the minor class was a high-mannose type (Man5GlcNAc2 (7.5%)). This is the first report to show that alpha(1-->3) fucosylation of N-glycans does occur but beta(1-->4) galactosylation of the sugar chains does not in the tobacco cultured cells.

The effect of pioglitazone on glucose metabolism and insulin uptake in the perfused liver and hindquarter of high-fructose-fed rats.

To investigate the effect of pioglitazone, a thiazolidinedione oral antidiabetic agent, on the glucose and insulin metabolism in insulin resistance, a perfusion study of the liver and hindquarter was performed in high-fructose-fed rats. Male Wistar albino rats were assigned randomly to one of the following diets for 2 weeks: (1) normal chow (control group), (2) a diet high in fructose (fructose group), or (3) a high-fructose diet plus pioglitazone (pioglitazone intake of approximately 10 mg/kg body weight; pioglitazone group). The elevated levels of plasma insulin, triglyceride, and free fatty acids (FFA) in the fructose group were normalized by pioglitazone administration. In the perfused liver, the glucagon-induced increment in the glucose output of the fructose (57.1+/-9.1 micromol/g liver/20 min) and pioglitazone (44.7+/-10.1 micromol/g liver/20 min) groups was significantly (P < .01) higher than that in the control group (27.6+/-5.7 micromol/g liver/20 min). The level in the pioglitazone group was significantly (P < .05) lower than that in the fructose group. In the presence of 100 or 500 microU/mL insulin, the insulin-mediated decrement in the glucagon-induced glucose output of the fructose group (29.8+/-7.8 or 38.9+/-9.3 micromol/g liver/20 min) was significantly (P < .05) lower than that in the control (45.8+/-14.2 or 54.5+/-8.5 micromol/g liver/20 min) and pioglitazone (44.4+/-9.2 or 56.2+/-10.8 micromol/g liver/20 min) groups, respectively. In the perfused hindquarter, glucose uptake in the fructose group (8.2+/-2.0 micromol/g muscle/30 min) was significantly (P < .05) lower than that in the control (12.1+/-2.3 micromol/g muscle/30 min) and pioglitazone (11.8+/-3.1 micromol/g muscle/30 min) groups. In the presence of 100 or 500 microU/mL insulin, glucose uptake in the fructose group (12.0+/-5.2 or 17.4+/-3.0 micromol/g muscle/30 min) was significantly (P < .05) lower than that in the control (20.2+/-2.4 or 23.0+/-3.1 micromol/g muscle/30 min) and pioglitazone (17.8+/-2.4 or 20.7+/-2.0 micromol/g muscle/30 min) groups, respectively. Insulin uptake by the liver and hindquarter was not significantly different in the control, fructose, and pioglitazone groups. These results indicate that pioglitazone improves the increased glucagon-induced hepatic glucose output and decreases insulin-induced muscular glucose uptake without altering insulin uptake in high-fructose-fed insulin-resistant rats.

[Bone changes in thyrotoxicosis].

Thyroid hormone (T3) is essential for normal bone growth and bone metabolism. T3 stimulates bone formation directly through T3 receptors in osteoblasts. T3 also stimulates bone resorption by osteoclasts probably secondary through osteoblasts. In thyrotoxicosis accelerated bone formation and resorption resulted in high turn-over bone loss. Bone metabolic markers elevate reflecting thyrotoxic state. Normalizing thyroid hormone level at least partially restore bone mineral content. In patients under thyroid hormone replacement therapy or TSH suppression therapy TSH and free thyroid hormones should be monitored to prevent unnecessary bone loss. Especially in postmenopausal women with thyrotoxicosis or thyroid hormone therapy the assessment of bone mineral content is required.