[Show abstract][Hide abstract] ABSTRACT: Insulin stimulation results in the activation of cyclin-dependent kinase-5 (CDK5) in lipid raft domains via a Fyn-dependent phosphorylation on tyrosine residue 15. In turn, activated CDK5 phosphorylates the Rho family GTP-binding protein TC10alpha on threonine 197 that is sensitive to the CDK5 inhibitor olomoucine and blocked by small interfering RNA-mediated knockdown of CDK5. The phosphorylation deficient mutant T197A-TC10alpha was not phosphorylated and excluded from the lipid raft domain, whereas the phosphorylation mimetic mutant (T197D-TC10alpha) was lipid raft localized. Insulin resulted in the GTP loading of T197D-TC10alpha but not T197A-TC10alpha and in parallel, T197D-TC10alpha but not T197A-TC10alpha depolymerized cortical actin and inhibited insulin-stimulated GLUT4 translocation. These data demonstrate that CDK5-dependent phosphorylation maintains TC10alpha in lipid raft compartments thereby disrupting cortical actin, whereas subsequent dephosphorylation of TC10alpha through inactivation of CDK5 allows for the re-assembly of F-actin. Because cortical actin reorganization is required for insulin-stimulated GLUT4 translocation, these data are consistent with a CDK5-dependent TC10alpha cycling between lipid raft and non-lipid raft compartments.
Journal of Biological Chemistry 11/2008; 283(51):35455-63. · 4.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The contribution of fasting and postprandial glucose to hemoglobin A(1c) (HbA(1c)) levels was evaluated in insulin-treated patients. In 57 insulin-treated, diabetic out-patients, fasting glucose (before breakfast (B-FG), lunch (L-FG) and dinner (D-FG)) and postprandial glucose (B-PPG, L-PPG and D-PPG) levels were determined by the patients themselves at home using glucose self-monitoring apparatus over the course of one week. The correlation between HbA(1c) levels and self monitored blood glucose levels were calculated. In the conventionally treated group, there was a significant correlation between HbA(1c) and fasting glucose (FG) levels only before lunch, but at 2 hr after (PPG) all meals. In the intensively treated group, a significant correlation between HbA(1c) levels and FG levels was found before lunch and at 2 hr after breakfast and dinner. In all subjects, only FG levels before lunch correlated significantly with HbA(1c) levels, although PPG levels were significantly correlated with HbA(1c) at all points. The correlation was highest with PPG after breakfast and dinner. The sum of all FG, PPG and FG + PPG levels was significantly correlated with HbA(1c) levels. Postprandial hyperglycemia after breakfast and dinner should be regarded as most important for improving HbA(1c) levels in insulin treated diabetic patients.
[Show abstract][Hide abstract] ABSTRACT: Previously we identified an unusual potential dual Akt/protein kinase B consensus phosphorylation motif in the protein Synip (RxKxRS(97)xS(99)) with serine 99 as a unique Akt2, but not Akt1 or for Akt3, substrate phosphorylation site. Although we have previously reported that serine 99 to phenylalanine (S99F-Synip) resulted in a constitutive inhibition of insulin-stimulated Glut4 translocation, a recent report indicated that Synip serine 99 to alanine mutant (S99A-Synip) failed to inhibit insulin-stimulated Glut4 translocation [H. Sano, S. Kane, E. Sano, G.E. Lienhard, Synip phosphorylation does not regulate insulin-stimulated GLUT4 translocation, Biochem. Biophys. Res. Commun. 332 (2005) 880-884]. To address this apparent discrepancy, we have now examined the S99A-Synip mutant and find that this mutant behaves essentially identical to S99F-Synip in that overexpression inhibits insulin-stimulated Glut4 translocation and is incapable of undergoing insulin-stimulated Syntaxin4 dissociation. These data are consistent with Synip serine 99 phosphorylation required for insulin-stimulated Glut4 translocation.
Biochemical and Biophysical Research Communications 05/2007; 356(1):102-6. · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: High-fat diets cause peripheral leptin resistance, and dietary lipid composition affects sensitivity to leptin. We examined the role of n-3 polyunsaturated fatty acid (PUFA) in peripheral leptin resistance. Dietary PUFAs (0.4% wt/wt) caused insensitivity to peripherally but not intracerebroventricularly administered leptin. n-3 PUFA increased body weight, associated with a significant reduction of leptin concentration in the cerebrospinal fluid. Dietary n-3 PUFA reduced transport of endogenous or exogenously administered leptin into the brain, associated with increased expression of hypothalamic occludin, but caused no change in expression of leptin receptors, proteins associated with leptin signaling or other tight junction proteins. Continuous intracerebroventricular infusion of an antisense morpholino oligonucleotide targeted to occludin mRNA reversed n-3 PUFA-induced insensitivity to peripherally administered leptin. We conclude that n-3 PUFA induces peripheral leptin resistance via an increase in the expression of hypothalamic occludin, reducing paracellular transport of leptin into the brain.
[Show abstract][Hide abstract] ABSTRACT: We have identified an unusual potential dual Akt/protein kinase B consensus phosphorylation motif in the protein Synip (RxKxRS(97)xS(99)). Surprisingly, serine 97 is not appreciably phosphorylated, whereas serine 99 is only a specific substrate for Akt2 but not Akt1 or Akt3. Although wild-type Synip (WT-Synip) undergoes an insulin-stimulated dissociation from Syntaxin4, the Synip serine 99 to phenylalanine mutant (S99F-Synip) is resistant to Akt2 phosphorylation and fails to display insulin-stimulated Syntaxin4 dissociation. Furthermore, overexpression of WT-Synip in 3T3L1 adipocytes had no effect on insulin-stimulated recruitment of glucose transporter 4 (GLUT4) to the plasma membrane, whereas overexpression of S99F-Synip functioned in a dominant-interfering manner by preventing insulin-stimulated GLUT4 recruitment and plasma membrane fusion. These data demonstrate that insulin activation of Akt2 specifically regulates the docking/fusion step of GLUT4-containing vesicles at the plasma membrane through the regulation of Synip phosphorylation and Synip-Syntaxin4 interaction.
The Journal of Cell Biology 04/2005; 168(6):921-8. · 9.69 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To determine the downstream signaling pathways regulated by betacellulin (BTC) in comparison with epidermal growth factor (EGF), we used Chinese hamster ovary cells overexpressing the human EGF receptor (ErbB1/EGFR). The overall time-dependent activation of EGFR autophosphorylation was identical in cells treated with 1 nm BTC or 1.5 nm EGF. Analysis of site-specific EGFR phosphorylation demonstrated that the BTC and EGF tyrosine phosphorylation of Y1086 was not significantly different. In contrast, the autophosphorylation of Y1173 was markedly reduced in BTC-stimulated cells, compared with EGF stimulation that directly correlated with a reduced BTC stimulation of Shc tyrosine phosphorylation, Ras, and Raf-1 activation. On the other hand, Y1068 phosphorylation was significantly increased after BTC stimulation, compared with EGF in parallel with a greater extent of Erk phosphorylation. Expression of a dominant interfering MEK kinase 1 (MEKK1) and Y1068F EGFR more efficiently blocked the enhanced Erk activation by BTC, compared with EGF. Interestingly BTC had a greater inhibitory effect on apoptosis, compared with EGF, and expression of Y1068F EGFR abolished this enhanced inhibitory effect. Together, these data indicated that although BTC and EGF share overlapping signaling properties, the ability of BTC to enhance Erk activation occurs independent of Ras. The increased BTC activation results from a greater extent of Y1068 EGFR tyrosine phosphorylation and subsequent increased recruitment of the Grb2-MEKK1 complex to the plasma membrane, compared with EGF stimulation. The increased Erk activation by BTC associated with antiapoptotic function.
[Show abstract][Hide abstract] ABSTRACT: A 24-year-old woman was admitted to our department for further examination of hypercalcemia, a high level of intact parathyroid hormone (PTH) and a right parathyroid tumor. She complained of bone pain throughout her body and was unable to walk due to systemic cystic osteofibrosis, including a brown tumor of the left lower extremities. Neck ultrasonography (US) and magnetic resonance imaging (MRI) revealed a tumor 2 cm in diameter in the upper side of the right thyroid lobe. 99mTc sestamibi (99mTc-MIBI) imaging and F-18 fluorodeoxyglucose positron emission tomography (FDG-PET) were performed to diagnose primary hyperparathyroidism and examination of other parathyroid glands. However, neither imaging modality detected the parathyroid tumor. To confirm the diagnosis, we performed selective venous sampling around the parathyroid and the patient was diagnosed with primary hyperparathyroidism due to a right parathyroid tumor. Resection of the right parathyroid tumor was performed and the pathological diagnosis was parathyroid adenoma. To date, both 99mTc-MIBI and FDG-PET are useful to localize parathyroid tumors. In this case, however, neither modality detected the tumor. Although recent studies state that expression of P-glycoprotein (P-gp) in parathyroid tumors plays an important role in the false-negative results of both 99mTc-MIBI scans and FDG-PET, immunohistological study detected no P-gp expression in the parathyroid tumor in the current case.
Internal Medicine 10/2004; 43(9):816-23. · 0.97 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We have measured the visible light spectrum emitted by hydrogen atoms adsorbed on an Ni(110) surface, excited by the tunneling current from the scanning tunneling microscope. The spectrum contains periodic fine structures, whose period corresponds to the vertical vibrational energy of the adsorbed H atom. This energy showed the expected isotope shift when H was replaced by deuterium, and further it depended on the H-atom chain length.
[Show abstract][Hide abstract] ABSTRACT: We have investigated the scanning-tunneling microscope (STM) light-emission mechanism of the Au(110)-(2×1) surface. We found that the light stimulated by the STM is emitted through three different channels. The first channel is the emission through excitation of localized surface plasmons (LSP). The other two channels are through the recombination of d-band holes and s-p electrons in Au. When the sample bias voltage is positive (i.e., electrons are injected into the sample), d-band holes are created by impact ionization. The intensity due to this process is greater when the tip is located between the Au atomic rows than over the row. This process is the origin of the atomic-site-dependent spectra that we reported in a previous paper [Y. Uehara, T. Fujita, and S. Ushioda, Phys. Rev. Lett. 83, 2445 (1999)]. When the bias voltage is negative, d-band holes are created by tunneling of d electrons from Au to the tip. The light-emission intensity due to the recombination of these d holes with sp electrons is about twice as strong as that emitted through the excitation of LSP.
Physical Review B 10/2002; 66(16). · 3.66 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Withdrawal of estrogen by ovariectomy increases adiposity, but decreases the circulating levels of the ob gene product, leptin, which inhibits food intake. The reduction of circulating leptin levels may thus play an important role in the induction of obesity by ovariectomy. To examine this hypothesis, body weight change by ovariectomy was investigated in leptin-deficient genetically obese (ob/ob) mice with leptin supplement. Prior to the operation, obese (ob/ob) female mice were treated with intraperitoneal administration of recombinant mouse leptin (1.0 microg/g body weight/day) for 8 days. Then, half of the leptin-treated mice and their lean littermates were bilaterally ovariectomized and their body weight changes were observed for 56 days. From 16 days after the operation, a significant increase in body weight by ovariectomy was observed only in lean mice without leptin treatment. From 44 days, a significant body weight gain by ovariectomy was observed in leptin-treated obese mice. Ovariectomy significantly increased retroperitoneal white adipose tissue weight in their lean littermates, but not in leptin-treated obese mice. It was suggested that the reduction of circulating leptin levels may play an important role in the increases of acute phase body weight gain by ovariectomy, but during static phase, the direct effects of estrogen withdrawal may appear independent of leptin-mediated effects.
[Show abstract][Hide abstract] ABSTRACT: We have investigated the Ni(110)–(2×1) O surface by scanning tunneling microscope (STM) light emission spectroscopy. A mosaic of light and dark domains was seen in the STM image of the surface. The STM light emission spectra of both domains were measured to compare with that of the clean Ni(110)–(1×1) surface. We found that the dark domains are covered by the O atoms and the light domains correspond to the exposed Ni surface.
Solid State Communications 06/2002; 122:451-453. · 1.70 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Concentrations of the soluble leptin receptor (sOB-R) may be related to leptin resistance in obesity. We measured sOB-R concentrations in serum in 103 non-diabetic Japanese men and women. All subjects were grouped according to body mass index (BMI; in kg/m(2)). Serum sOB-R concentrations did not differ significantly between normal-weight (18.5 < or = BMI < 25.0) men and women, but were significantly higher in underweight subjects (BMI < 18.5) than in normal-weight subjects. In contrast, overweight (25 < or = BMI < 30) and obese (30 < or = BMI < 35.0, 35.0 < or = BMI < 40, and BMI > or = 40) subjects had significantly lower sOB-R concentrations than did normal-weight subjects. Serum sOB-R concentrations were inversely correlated with BMI and serum immunoreactive leptin concentrations. Very low-energy diet therapy for 4 wk significantly lowered serum immunoreactive leptin concentrations but did not significantly affect serum sOB-R concentrations. Serum sOB-R concentrations did not change significantly during the menstrual cycle. Our results showed that serum sOB-R concentrations decrease with increasing BMI and that sex hormones likely do not affect serum sOB-R concentrations in non-pregnant women. The reduction in serum sOB-R concentrations in overweight and obese persons may reflect downregulation of hypothalamic leptin receptor production as a result of an increase in circulating leptin and might be an important factor in leptin resistance.
[Show abstract][Hide abstract] ABSTRACT: We have identified electronic transitions tightly localized at a single oxygen atom adsorbed on the Cu(110)-(2×1) surface, by means of scanning tunneling microscope (STM) light emission spectroscopy. The emission spectrum measured with the STM tip located directly over the oxygen atom shows two peaks corresponding to states located ∼200 and ∼300 meV above the Fermi level of Cu. These peaks are identified with unoccupied states localized at a single oxygen atom.
Solid State Communications 09/2001; 119(12-119):671-674. · 1.70 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There appears to be an obvious difference in the susceptibility to diabetic microangiopathy among Type 2 diabetic (T2DM) patients. We examined the hypothesis that aldose reductase (AR) messenger ribonucleic acid (mRNA) expression may be enhanced in diabetic patients susceptible to diabetic microangiopathy by measuring AR mRNA expression in the polymorphonuclear leukocytes (PMNs) obtained from T2DM patients. The AR mRNA and beta-actin mRNA expressions were measured by reverse transcription-polymerase chain reaction method. The relative AR mRNA expression to beta-actin mRNA expression significantly increased in the PMNs of T2DM patients, compared to normal healthy controls. The relative AR mRNA expression tended to increase with an increase of glycosylated haemoglobin A1c (HbA1c) levels. The patients who showed both progressive diabetic retinopathy and diabetic neuropathy within 10 years after the diagnosis of T2DM (Group I) showed significantly 1.6 times higher AR mRNA expressions than those who have not shown both complications over 10 years after diagnosis (Group II), in spite of no obvious difference in glycaemic control between both groups. The present data indicate that the AR mRNA expression increased in the PMNs of T2DM patients and there is an obvious difference in the PMNs AR mRNA expression between T2DM patients susceptible and relatively resistant to the development of diabetic microangiopathy.
[Show abstract][Hide abstract] ABSTRACT: Leptin, ob gene product, inhibits feeding behavior and stimulates energy expenditure in rodents. Corticotropin-releasing hormone (CRH) and neuropeptide Y (NPY), which act in the hypothalamus to influence energy homeostasis, may mediate the anorexic effect of leptin. The present studies were undertaken to examine the possible involvement of hypothalamic CRH in the anorexigenic action of leptin in male Wistar rats. Recombinant leptin (2 microg/rat), microinjected into the third ventricle, inhibited food intake at 2 h by 33.3% (P < 0.01) in rats that were deprived of food for 18 h. The intracerebroventricular injection of 2 microg leptin also increased hypothalamic CRH content (P < 0.05) at 2 h after its administration. Simultaneous intracerebroventricular administration of 5 microg/rat alpha-helical CRH 9-41 (alpha-hCRH), a CRH antagonist, with 2 microg/rat leptin attenuated the anorexic effect of leptin by 2 h. In contrast, single intracerebroventricular injection of alpha-hCRH did not affect food consumption in food-deprived rats. These results implicate hypothalamic CRH as an important mediator of the anorexic effect of leptin in food-deprived rats.