Effect of Gadolinium on the Estimation of Myelin and Brain Tissue Volumes Based on Quantitative Synthetic MRI.

BACKGROUND AND PURPOSE: The effect of gadolinium on the estimation of myelin has not been reported. The aim of the current study was to investigate the effects of gadolinium on automatic myelin and brain tissue volumetry via quantitative synthetic MR imaging. MATERIALS AND METHODS: The study included 36 patients who were referred for brain metastases screening, and quantitative synthetic MR imaging data before and after gadolinium-based contrast agent administration were analyzed retrospectively. Brain metastases were detected in 17 patients. WM volume, GM volume, CSF volume, non-WM/GM/CSF volume, myelin volume, brain parenchymal volume, myelin fraction (myelin volume/brain parenchymal volume), and intracranial volume were estimated. T1 and T2 relaxation times, proton density, and myelin partial volume per voxel averaged across the brain parenchyma were also analyzed. RESULTS: In patients with and without metastases after gadolinium-based contrast agent administration, measurements of WM and myelin volumes, and myelin fraction were significantly increased (+26.65 and +29.42 mL, +10.14 and +12.46 mL, +0.88% and +1.09%, respectively), whereas measurements of GM, CSF, brain parenchymal, and intracranial volumes were significantly decreased (-36.23 and -34.49 mL, -20.77 and -18.94 mL, -6.76 and -2.84 mL, -27.41 and -21.84 mL, respectively). Non-WM/GM/CSF volume did not show a significant change. T1, T2, and proton density were significantly decreased (-51.34 and -46.84 ms, -2.67 and -4.70 ms, -1.05%, and -1.28%, respectively) after gadolinium-based contrast agent administration, whereas measurements of myelin partial volume were significantly increased (+0.78% and +0.75%, respectively). CONCLUSIONS: Gadolinium had a significant effect on the automatic calculation of myelin and brain tissue volumes using quantitative synthetic MR imaging, which can be explained by decreases in T1, T2, and proton density.

New constraint on the existence of the µ+ → e+ γ decay.

The analysis of a combined data set, totaling 3.6 × 10(14) stopped muons on target, in the search for the lepton flavor violating decay µ(+) → e(+)γ is presented. The data collected by the MEG experiment at the Paul Scherrer Institut show no excess of events compared to background expectations and yield a new upper limit on the branching ratio of this decay of 5.7 × 10(-13) (90% confidence level). This represents a four times more stringent limit than the previous world best limit set by MEG.

Three-dimensional images of liver tumours reconstructed by Gd-EOB-DTPA-enhanced MRI.

The purpose of this study was to evaluate three-dimensional images of liver tumours obtained with gadolinium ethoxybenzyl diethylenetriamine penta-acetic acid (Gd-EOB-DTPA)-enhanced MRI (3D-EOB-MRI) in hepatic surgery. We conclude that 3D-EOB-MRI may be an alternative method for depicting liver tumours adjacent to the hepatic veins and portal branches, and may provide additional information for surgical planning.

New limit on the lepton-flavor-violating decay µ+→e+γ.

We present a new result based on an analysis of the data collected by the MEG detector at the Paul Scherrer Institut in 2009 and 2010, in search of the lepton-flavor-violating decay µ(+)e(+)γ. The likelihood analysis of the combined data sample, which corresponds to a total of 1.8×10(14) muon decays, gives a 90% C.L. upper limit of 2.4×10(-12) on the branching ratio of the µ(+)→e(+)γ decay, constituting the most stringent limit on the existence of this decay to date.

Imaging of early pancreatic cancer on multidetector row helical computed tomography.

Early pancreatic cancer is small and limited to the pancreas. In contrast, small pancreatic cancer may include peripancreatic vasculature or metastasis involvement. This study evaluates images of early pancreatic cancer on multidetector CT (MDCT) using contrast-enhanced multiphasic imaging, and post-processed pancreatic duct images. CT findings and pathological features were analysed in eight patients with early pancreatic cancer. Pathological evaluation included location, size and histological grading of the tumour. MDCT evaluation covered the maximum diameter of the main pancreatic duct (MPD), stenosis or obstruction of the MPD, loss of normal lobar texture and associated pancreatitis. Attenuation differences between normal pancreatic parenchyma and the tumour (AD-PT) were also measured. Focal stenosis or obstruction of the MPD with dilatation of the distal MPD was demonstrated in all patients. Associated pancreatitis occurred in six patients with tumours measuring 12 mm or greater. Loss of normal lobar texture was recognised in four cases with the tumour measuring 14 mm or greater. Statistically, low-attenuated lesions and high-attenuated lesions differed with respect to the tumour size (p<0.01), and a positive relationship was demonstrated between the tumour size and AD-PT (r = 0.84). In seven cases, AD-PT is higher during the arterial phase than the pancreatic phase. Early pancreatic cancer appears as low attenuation on early phase, and as high- to iso-attenuation during the pancreatic and delayed phases in respect to the tumour size. Focal stenosis or obstruction of the MPD with dilatation of the distal MPD observed on curved reformation imaging seems important in the diagnosis of early pancreatic cancer.

Direct production of an activated matrix metalloproteinase-9 (gelatinase B) from mammalian cells.

Matrix metalloproteinase-9 (MMP-9) is produced by the inactive proform and activated by a proteolytic process. However, it has not been reported to produce the active form directly from cells, which has hindered the research to elicit the physiological roles of this enzyme. In this study, we prepared mutant MMP-9 containing the furin-recognizing sequence in the prodomain and showed that the mutant MMP-9 was secreted as the active form directly from CHO-K1 cells and primary hepatocytes after the gene was transfected. The secreted MMP-9 showed proteolytic activity without further activation and degraded collagen IV in vitro. In addition, the transfection of the gene into the liver resulted in the efficient expression of active MMP-9 in the liver and the serum in vivo.

Osteogenesis coordinated in C3H10T1/2 cells by adipogenesis-dependent BMP-2 expression system.

A novel tissue engineering for bone formation has been proposed, to make osteoblast differentiation balanced by transfecting the mesenchymal stem cells with a gene encoding human bone morphogenetic protein-2 (hBMP-2) under the control of adipocyte specific lipoprotein lipase (LPL) promoter. Due to the promoter specificity, the initiation of BMP transcription is dependent on adipogenesis. For 14-day culture in the presence of ascorbic acid (asc) and beta-glycerophosphate (gly), nontransfected mouse embryonic fibroblast C3H10T1/2 (10T1/2) cells showed extensive accumulation of lipid droplets and adipocyte specific enzyme glycerol-3-phosphate dehydrogenase (G3PDH) mRNA expression, but exhibited neither BMP-2 expression, high alkaline phosphatase (ALP) activity which reflects osteoblast phenotype. On the other hand, transfected 10T1/2 cells showed hBMP-2 expression, high ALP activity and low level of G3PDH. mRNA expression accompanied with minimal lipid droplets. These results indicate that 10T1/2 cells are proved to be differentiated with maintaining coordinated balance of adipogenesis and osteogenesis, when they are transfected by the gene encoding hBMP-2 under the control of LPL promoter.

Regulation and significance of hepatocyte-derived matrix metalloproteinases in liver remodeling.

Regulation in expression and activation of proteinases is one of the most important mechanisms in organ morphogenesis. In this study, we investigated the expression of MMPs in primary hepatocytes and their roles in liver remodeling. A hepatocyte proliferation initiating cytokine, TNFalpha, induced MMP-9 expression in these cells while the expression of MMP-2 did not change by zymography analysis. Interestingly, both the induced MMP-9 expression and hepatocyte proliferation by TNFalpha were synergistically enhanced by HGF in vitro. The increased proliferation was suppressed by MMP inhibitor TIMP-1, suggesting that cytokine-induced MMP regulates proliferation. The increased expression of MMP-9 by the cytokines was inhibited by cytochalasin D or colchicine but not by PI3 kinase inhibitor wortmannin. In addition, co-stimulation by TNFalpha and HGF of spheroidal hepatocytes cultured in 3-dimensional collagen gel drastically induced morphological changes by cell extension and migration in the gel, which was in parallel with the induced expression of MMP-9 and was inhibited by TIMP-1 and -2. The MMP activity was also detected in vivo in the regenerating liver after partial hepatectomy by in situ zymography. These results suggest the roles of MMPs produced by parenchymal cells in liver remodeling.

Assembling of engineered IgG-binding protein on gold surface for highly oriented antibody immobilization.

The B-domain, which is one of IgG-binding domains of staphylococcal protein A, was repeated five times and a cysteine residue was introduced at its C-terminus by a genetic engineering technique. The resulting protein, designated B5C1, retained the same IgG-binding activity as native protein A. The B5C1 was assembled on a gold plate surface by utilizing a strong affinity between thiol of cysteine and a gold surface. IgG-binding activity of B5C1 on a gold surface was much higher than that of physically adsorbed B5, which lacks cysteine residue. Furthermore, antigen-binding activity of immobilized antibody molecules through the use of assembled B5C1 on a gold surface was about 4.3 times higher than that of physically adsorbed antibody molecules. Immobilization of highly oriented antibody molecules was realized with the engineered IgG-binding protein.

Presence of telomeric G-strand tails in the telomerase catalytic subunit TERT knockout mice.

BACKGROUND: Telomerase consists of two essential subunits, the template RNA (TR; telomerase RNA) and the catalytic subunit TERT (telomerase reverse transcriptase). Knockout mice with a mTR (mouse TR) deletion have been described and well characterized. However, mice with a mTERT (mouse TERT) deletion have not been reported. RESULTS: mTERT-knockout mice have been constructed. The first generation mTERT -/- mice were fertile, and did not show any noticeable macroscopic or microscopic phenotypic change. All tissue cells derived from mTERT -/- mice that were examined lacked telomerase activity, indicating that mTERT is the only gene encoding the telomerase catalytic subunit. Pulse field gel electrophoresis (PFGE) and nondenaturing in-gel hybridization analyses showed that mouse telomeric DNA has G-strand 5'-overhangs, as demonstrated for human and yeast cells. This telomeric single-stranded G-tail was also observed in MEF (mouse embryonic fibroblast) and liver cells derived from mTERT -/- mice. CONCLUSIONS: mTERT-knockout mice show phenotypes that are apparently normal at least during the early generations. This observation is similar to that obtained with the mTR-knockout mice. The presence of the telomeric G-strand tails in mTERT -/- mice suggests that these telomeric 5'-overhangs are produced by telomerase-independent mechanisms, as has been proposed for yeast and human.

Site-directed lipid modification of IgG-binding protein by intracellular bacterial lipoprotein process.

IgG-binding protein was genetically expressed and lipid-modified in a site-directed manner in Escherichia coli. The DNA sequence encoding the signal peptide and the nine N-terminal amino acid residues of the major lipoprotein of E. coli (lpp) was fused to the sequence of B-domain which was one of the IgG binding domains of Staphylococcal Protein A (SpA). The N-terminal cysteine residue of the resulting protein was enzymatically linked with lipids in the bacterial membrane. The lipid-modified protein was translocated at the bacterial membrane in a manner similar to native bacterial lipoprotein, and it was purified with IgG-Sepharose by affinity chromatography. The lipid modified proteins (lppB1 and lppB5) showed a similar IgG binding activity to unmodified proteins, which was estimated by competitive ELISA. Proteoliposomes of lipid modified proteins were prepared in an elegant fashion so that the IgG binding site should be properly oriented on the surface of an individual liposome by anchoring the lipid-tail into the hydrophobic layer of the liposome membrane. As compared with the unmodified one, the lipid modified protein incorporated into the proteoliposome exhibited higher IgG binding activity.

IRF-1 is an essential mediator in IFN-gamma-induced cell cycle arrest and apoptosis of primary cultured hepatocytes.

IFN-gamma induces cell cycle arrest and p53-independent apoptosis in primary cultured hepatocytes. However, it is not yet understood what molecules regulate the mechanism. We report here that interferon regulatory factor 1 (IRF-1) is an essential molecule in these phenomena. Hepatocytes from IRF-1-deficient mice were completely resistant to IFN-gamma in apoptosis indicated by three different hallmarks such as LDH release, DNA fragmentation and the activation of caspase-3 family. Caspase-1 expression was little detected in hepatocytes, and constitutive and IFN-gamma-induced mRNA expression of Fas or caspase-3 did not change in between wild type and IRF-1-deficient hepatocytes. Expression of IFN-gamma-inducible caspase, caspase-11, did not change either. Thus, it is unlikely that these molecules directly regulate the mechanisms. Interestingly, IRF-1-deficient hepatocytes were also resistant to IFN-gamma-induced cell cycle arrest despite IFN-gamma-induced cell cycle arrest and apoptosis are regulated by independent pathways. Results by Northern blot analysis showed that IFN-gamma-induced but not constitutive p53 mRNA expression was regulated by IRF-1. In fact, IFN-gamma did not induce cell cycle arrest in p53-deficient hepatocytes. Taken together, IRF-1 mediates IFN-gamma signaling into primary hepatocytes for cell cycle arrest via p53 expression and for apoptosis.

Two types of electrochemical nitric oxide (NO) sensing systems with heat-denatured Cyt C and radical scavenger PTIO.

To develop an in situ NO sensing system for primarily biological and medical uses, two types of NO sensing materials, which may be coupled with an electrochemical reaction for signal transduction, have been investigated. Heat-denatured Cyt C and a radical scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetra methyl imidazoline-1-oxyl-3-oxide (C-PTIO) were found to be effective and were incorporated into electrochemical sensing systems. Heat-denatured Cyt C deposited on a 4-mercaptopyridine modified gold electrode responded to NO with an increase of cathodic current through electrochemical reduction of Cyt C (Fe3+), when the electrode potential was controlled at 0 mV vs Ag/AgCl. The dynamic range of the sensing system was 0.5-4 microM. The sensing system with C-PTIO exhibited an anodic output in response to NO at 0.7 V vs Ag/AgCl, and showed a wider dynamic range from 0.05 to 100 microM.

Gene expression in the electrically stimulated differentiation of PC12 cells.

Cell differentiation of PC12 cells was electrically induced to grow neurites in the absence of nerve growth factor (NGF) on the electrode surface, of which potential was modulated by a rectangular wave of potential. The electric stimulation induced the c-fos expression which is essential for cell differentiation. Non-specific calcium channel blocker, lanthanum ion, inhibited the electrically induced differentiation, while NGF-induced differentiation was not suppressed. An L-type calcium channel blocker, nifedipine, also inhibited the electrically induced calcium influx and c-fos expression. Moreover, a stretch-activated (SA) channel blocker, gadolinium ion, inhibited the electrically stimulated differentiation by blocking the calcium influx, but gave no prominent effects on the potassium ion-induced differentiation. Chelerythrine, a specific protein kinase C (PKC) inhibitor, almost inhibited the cell differentiation by the electric stimulation but not by the NGF treatment. These results indicate that the alternative potential may stimulate cell differentiation through a PKC cascade.

Use of a quartz crystal microbalance to monitor immunoliposome--antigen interaction.

We have used quartz crystal microbalance (QCM)-based real-time biospecific interaction measurement to analyze the binding of immunoliposomes to antigen and examined the use of liposomes as signal-enhancing reagents in competitive QCM immunoassay. For the preparation of immunoliposomes, various amounts of bacterially produced lipid-tagged single-chain antibody against 2-phenyloxazolone were incorporated in phosphatidylcholine liposomes. The immunoliposomes bound specifically to immobilized hapten, and this binding was inhibited by soluble hapten in a concentration-dependent manner. In this competitive assay, antigen could be measured in the concentration range from 10(-5) to 10(-8) M.

Electrically induced neurite outgrowth of PC12 cells on the electrode surface.

Morphological differentiation of PC12 cells cultured on an indium-tin oxide (ITO) electrode has been induced to grow neurites in the absence of nerve growth factor (NGF) by electrical stimulation. Rectangular pulse wave potentials were applied to the electrode at amplitudes of 200 mV and 400 mV with frequencies of 50 Hz, 500 Hz, and 1 kHz. The PC12 cells differentiated most prominently at 200 mV with 100 Hz. No statistically significant differences were observed among the electrically induced neurite lengths. The electrically induced differentiation was completely inhibited by a blockade of calcium influx using LaCl3. This indicates that repeated potential shift in the vicinity of a cellular membrane may stimulate morphological response, probably through calcium ion channels.

A fluoroimmunoassay based on immunoliposomes containing genetically engineered lipid-tagged antibody.

Immunoliposomes were prepared by using biosynthetically lipid-tagged anti-2-phenyloxazolone single-chain antibody. Carboxyfluorescein as a fluorescent marker was encapsulated in the immunoliposomes. Some conditions for fluoroimmunoassay using the immunoliposomes were optimized by binding assays with hapten-coated microtiter wells. A competitive fluoroimmunoassay for the caproic acid conjugate of 2-phenyloxazolone as a model antigen was performed with the immunoliposomes. In the optimized assay conditions, antigen could be determined in the concentration range from 10(-7) to 10(-9) M.

Electrically induced NGF production by astroglial cells.

A controlled culture system has been developed to induce nerve growth factor (NGF) production in astroglial cells that are cultured on an electrode surface. The electrode potential is alternatively modulated at an amplitude of 300 mV and a frequency of 10 Hz. The electric stimulation triggers NGF production and secretion. The mechanism of the electrically induced NGF production is discussed in association with protein kinase C (PKC) activation.

On-off switching of enzymatic reaction by recombinant calmodulin on a solid-phase matrix.

A fusion protein consisting of human calmodulin (CaM) and glutathione S-transferase (GST) was produced by gene fusion. The fusion protein was overexpressed in Escherichia coli as a soluble form and purified with one-step affinity chromatography using glutathione-Sepharose. The protein had the modulating activity of CaM and the binding capability to glutathione of GST. Phosphodiesterase, which is a CaM dependent enzyme, was activated by the fusion protein, with the Ca2+ level equal to the level equivalent to a native CaM. Furthermore, CaM could be immobilized on a solid-phase matrix through the use of GST moiety while its modulating activity was retained. Phosphodiesterase activity was switched on and off by the immobilized CaM with or without Ca2+, and repeated use of CaM was demonstrated.

[A case of Addison's disease complicated by lung carcinoma treated with surgical resection--a case report].

Surgery was successful in a 72-year-old man with Addison's disease complicated by right lower lobe carcinoma of the lung. The patient had been diagnosed Addison's disease 25 years earlier and had been treated with cortisone acetate 25 mg every morning. A tumor shadow was found in the right lower lobe of the lung on chest X-ray. Squamous cell carcinoma was diagnosed by cytological examination. Right lower lobectomy with partial resection of the right diaphragma and lymph-node dissection were performed. The bronchial stump was closed using Sweet's method and covered with a pericardial fat pad. The intra- and post-operative course was uneventful with intravenous administration of adequate doses of hydrocortisone. The patient has been free from the carcinoma for two years.