Effect of Reveromycin A on experimental tooth movement in OPG-/- mice.

In osteoprotegerin-deficient (OPG-/-) mice, osteoclast activity causes bone resorption to outpace bone formation, leading to the development of severe osteoporosis. Such mice are therefore useful for investigating the alveolar bone of patients with osteoporosis. Reveromycin A (RM-A) was recently identified as the unique agent acting on osteoclast activation. This study aimed to analyze the effect of RM-A on the orthodontic treatment of OPG-/- mice (a model of osteoporosis patients with high levels of bone turnover). We examined alveolar bone remodeling in OPG-/- and wild-type (WT) mice during continuous tooth movement. The orthodontic force was induced by means of a Ni-Ti closed-coil spring to move the maxillary first molar for 14 days. RM-A sodium salt (1 mg/kg) was administered intraperitoneally twice daily. In OPG-/- mice, the tooth movement distance was longer, alveolar bone resorption was enhanced, the osteoclast count was greater, and serum alkaline phosphatase and tartrate-resistant acid phosphatase levels were higher relative to those in WT mice. However, the administration of RM-A in OPG-/- mice reduced these parameters. We conclude that RM-A normalizes bone metabolism and loss of alveolar bone during continuous tooth movement in OPG-/- mice.

Neurotoxic effects of methamphetamine on rat hippocampus pyramidal neurons.

Methamphetamine (MAP) is known to alter behavior and cause deficits in learning and memory. While the major site of action of MAP is on mesolimbic dopaminergic pathways, the effects on learning and memory raise the possibility of important actions in the hippocampus. We have studied electrophysiologic and morphologic effects of MAP in the CA1 region of hippocampus from young male rats chronically exposed to MAP, male rats exposed during gestation only and the effects of bath perfusion of MAP onto brain slices from control rats. Pyramidal neurons in brain slices from chronically exposed rats had reduced membrane potential and membrane resistance. Long-term potentiation (LTP) was reduced as compared to control, but when MAP was acutely perfused over control slices the amplitude of LTP was increased. LTP in young adult animals that had been gestationally exposed to MAP showed reduced LTP as compared to controls. Morphologically CA1 pyramidal neurons in chronically exposed animals showed a high prevalence of extensive blebbing of dendrites. We conclude that the NMDA receptor and the process of LTP are also targets of MAP dysfunction, at least in the hippocampus.

Proteomic signatures and aberrations of mouse embryonic stem cells containing a single human chromosome 21 in neuronal differentiation: an in vitro model of Down syndrome.

Neurodegeneration in fetal development of Down syndrome (DS) patients is proposed to result in apparent neuropathological abnormalities and to contribute to the phenotypic characteristics of mental retardation and premature development of Alzheimer disease. In order to identify the aberrant and specific genes involved in the early differentiation of DS neurons, we have utilized an in vitro neuronal differentiation system of mouse ES cells containing a single human chromosome 21 (TT2F/hChr21) with TT2F parental ES cells as a control. The paired protein extracts from TT2F and TT2F/hChr21 cells at several stages of neuronal differentiation were subjected to two-dimensional polyacrylamide gel electrophoresis protein separation followed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry to identify the proteins differentially expressed between TT2F and TT2F/hChr21 cells. We provide here a novel set of specific gene products altered in early differentiating DS neuronal cells, which differs from that identified in adult or fetal brain with DS. The aberrant protein expression in early differentiating neurons, due to the hChr21 gene dosage effects or chromosomal imbalance, may affect neuronal outgrowth, proliferation and differentiation, producing developmental abnormalities in neural patterning, which eventually leads to formation of a suboptimal functioning neuronal network in DS.

[Assessment of extracellular fluid in cardiac surgery using cardiopulmonary bypass].

Extracellular fluid (ECF) was assessed before and after the cardiac surgery using cardiopulmonary bypass (CPB), by means of a bioimpedance spectrum analyzer to see volumes of the fluid based on changes of the impedance to various frequencies. Difference between the levels before and after the operation was divided by body weight to study about a % BW. Simultaneously its relation to the lung compliance [tidal volume/(peak inspiratory pressure-end expiratory pressure)] was studied. Mean age of the 18 patients was 59.1 +/- 19 years old. ECF was assessed before to 24 hours after the operation continuously and once more after 48 hours. Mean CPB time was 165 +/- 52 minutes, and aortic cross clamp time was 121 +/- 4 minutes. A remarkable increase of ECF was noted immediately after the operation, which further increased gradually till arriving at the peak 4 hours after the operation (4.52 +/- 1.8% BW). Then it gradually decreased to 0.641 +/- 2.7% BW 48 hours later. Lung compliance measured at the same time showed the lowest level 6 hours after the operation. It was known that the bioimpedance spectrum analysis is a simple and non-invasive method, which enables to monitor the vital stable before and after the operation.

[Surgical treatment for giant ascending aorta-arch aneurysm coexisted with DeBakey type II dissection on elderly women: report of a case].

A 78-year-old woman was admitted to our hospital after computed tomography (CT) had revealed in her the presence of a giant ascending aorta-arch aneurysm. This aneurysm was about 8 cm in diameter and associated with DeBakey type II dissection. Aortography showed the same condition as the CT view with the entry on the ascending aorta. The ascending aortaarch was replaced with a Hemashield 24 mm, by using deep-hypothermic selective cerebral perfusion and the open distal method. There were no complications during her peripostoperative state and no evidence of leakage and remnant dissection on CT and aortography. This is a rare case in which thoracic aortic aneurysm coexisted with dissection. In this case of severe atherosclerosis, deep-hypothermic selective cerebral perfusion and the open distal method provided effective treatment.

A new mouse model for Down syndrome.

Trisomy 21 (Ts21) is the most common live-born human aneuploidy and results in a constellation of features known as Down syndrome (DS). Ts21 is a frequent cause of congenital heart defects and the leading genetic cause of mental retardation. Although overexpression of a gene(s) or gene cluster on human chromosome 21 (Chr 21) or the genome imbalance by Ts21 has been suggested to play a key role in bringing about the diverse DS phenotypes, little is known about the molecular mechanisms underlying the various phenotypes associated with DS. Four approaches have been used to model DS to investigate the gene dosage effects of an extra copy of Chr 21 on various phenotypes; 1) Transgenic mice overexpressing a single gene from Chr 21, 2) YAC/BAC/PAC transgenic mice containing a single gene or genes on Chr 21, 3) Mice with intact/partial trisomy 16, a region with homology to human Chr 21 and 4) Human Chr 21 transchromosomal (Tc) mice. Here we review our new model system for the study of DS using the Tc technology, including the biological effects of an additional Chr 21 in vivo and in vitro.

[A case of emergency coronary artery bypass graft for the patient using chronic hemodialysis, where in blood volume was measurable in the perioperative period].

An emergency coronary artery bypass graft (CABG) was given to a 66 year-old patient due to acute myocardial infarction (AMI). Circulating blood volume (BV) was measured to study in the perioperative period. Three coronary artery bypasses were made under cardiopulmonary bypass, being managed by ultrafiltration when the pump-oxygenator was in action and by peritoneal dialysis in the early postoperative period. Preoperative BV reduced immediately after the operation. It showed an increasing trend 4 hours after the operation, but after that BV reduced from that before the operation while water balance was kept positive. Cardiac output after the operation was higher than before. It suggested that in this patient using hemodialysis BV levels turned to be lower compared with that before the operation, as excessive water leaked out of the blood vessel, although water balance was kept positive due to improved cardiac functions after the operation.

[Blood transfusion under cardiopulmonary bypass is a possible inducer for inflammation?].

To investigate that blood transfusion under cardiopulmonary bypass is a possible inducer for inflammation, a retrospective study was made with 20 adult patients who underwent coronary artery bypass grafting. The subjects were divided into two groups; transfusion group (group T) including 9 patients who received blood transfusion during cardiopulmonary bypass and the control group (group C) including 11 patients who did not undergo perioperative transfusion. Respiratory index as an indicator of respiratory functions was determined before and immediately after cardiopulmonary bypass, at the end of surgery and 4 hours thereafter. Cardiac index and arterial pressure were determined as the indicator of cardiac function. Moreover, interleukin 6 and 8 (IL-6 and IL-8), inflammatory cytokines were measured and compared between the two groups. The mean amount of blood transfusion was 2.1 units per individual of group T. The minimum value of hematocrit during cardiopulmonary bypass was significantly lower in group T (15.8 +/- 1.8%) than group C (19.1 +/- 1.4%), but the difference became not significant after cardiopulmonary bypass. There were no significant differences either in aortic pressure or cardiac index between two groups. The respiratory index at the end of surgery was higher in group T but the difference was not significant. Meanwhile IL-8 level at the end of cardiopulmonary bypass was significantly higher in group T (67.9 +/- 36 pg/ml) than group C (35.1 +/- 21 pg/ml). However, there was no difference in IL-6 level between the two. These results suggested that inflammation might be aggravated by an increase of IL-8 induced by blood transfusion.

A method to maintain introduced DNA sequences stably and safely on the bacterial chromosome: application of prophage integration and subsequent designed excision.

By application of prophage integration and subsequent intended excision, a method to maintain an introduced DNA sequence stably onto a bacterial chromosome has been proposed. Recently-constructed integration plasmids using Campbell-type prophage integration system in Lactobacillus casei strain Shirota and its temperate phage phi FSW was modified for this purpose and a chloramphenicol (Cm)-resistance gene was used as a model passenger DNA. On the integration plasmid having an erythromycin (Em)-resistance gene as a selection marker, N- and C-terminally-truncated Cm-resistance genes were inserted into both sides of the attP of phi FSW, within which the site-specific recombination took place with the attB of phi FSW on the recipient chromosome through the phi FSW integrase. Primary integrants of the modified plasmid (integration-excision vector) exhibiting Em-resistant and Cm-sensitive phenotype generated Em-sensitive and Cm-resistant derivatives under the nonselective conditions. Sequence analyses showed that one copy of the complete Cm-resistance gene resided at the attachment site on the host chromosome and the other vector-derived sequences were excised probably by endogenous homologous recombination in the host cells to derive final integrants. The Cm-resistant phenotype of the final integrants was stable for more than 50 generations under non-selective conditions. Frequency of the homologous recombination suggests that negative selection is also adoptable. Thus, this method using the integration-excision vector gives a stable and safe derivatives of the strain and is likely to be applicable to various bacteria, since Campbell-type prophage integration system and homologous recombination are prevalent among bacteria.

Assessment of lung volumes in pulmonary emphysema using multidetector helical CT: comparison with pulmonary function tests.

The purpose of this study is to evaluate the usefulness of multidetector helical CT (MDCT) with three-dimensional (3D) postprocessing for assessing the lung volume at inspiration and expiration of the pulmonary emphysema and for comparing it with pulmonary function tests. Percentage lung volume at the threshold of -930, -900, -810, -790, and -770 at expiration showed good correlation with FEV1, FEV1/FVC, and DLCO/Va. Excellent correlation was observed between percentage lung volume at the threshold of -900 and FEV1/FVC. CT densitometry at expiration showed better correlation than that at inspiration with pulmonary function tests. MDCT with 3D technique is useful for assessing the severity of pulmonary emphysema.

Vascular encasement by pancreatic cancer: correlation of CT findings with surgical and pathologic results.

PURPOSE: The purpose of this study was to correlate thin-slice high-resolution helical CT findings of arterial and venous involvement in pancreatic cancers with surgical and histopathologic results. METHOD: Forty-eight patients with pancreatic cancer underwent preoperative thin-slice high-resolution helical CT, followed by surgical dissection of the pancreatic vessels during curative or palliative surgery. Major vessels running within 1 cm from the tumor margin were evaluated. CT appearance was graded on a 0-4 scale (0: none, 1: <24%, 2: 25-49%, 3: 50-74%, 4: 75-100%) by circumferential contiguity of tumor to vessels. Resected specimens were available from 26 patients. RESULTS: Surgical correlation of CT findings was available in 89 veins and 83 arteries, and both surgical and histologic correlation was available for 42 veins and 29 arteries. At surgical observation, 29 of 35 veins (82.9%) evaluated as CT grade 3 or 4 were found to be involved, whereas only 18 of 30 arteries (60%) evaluated as CT grade 3 or 4 were proved to be involved. On microscopic observation, tumor invasion to the portal venous systems was confirmed in 15 of 42 (35.7%) vessels, and this invasion was depicted as from CT grades 1 to 4. In arteries, tumor invasion was seen in 3 of 29 vessels (10.3%), all of which were graded as 3 or 4 by CT. CONCLUSION: The grading system of vascular invasion should differ between arteries and veins. Involvement of the venous system exceeding one-half circumference of the vessels (grade 3 or 4) was suggestive of vascular invasion; however, this criterion was not always satisfactory for the evaluation of tumor invasion in the arterial system.

Construction of 700 human/mouse A9 monochromosomal hybrids and analysis of imprinted genes on human chromosome 6.

As an in vitro assay system for the identification of human imprinted genes, a library of human/mouse A9 monochromosomal hybrids containing a single, intact bsr-tagged human chromosome of known parental origin, derived from normal human fibroblasts, has been previously generated by microcell-mediated chromosome transfer (MMCT). To supplement this assay system, we constructed additional 700 A9 monochromosomal hybrids, using a pSTneo or pPGKneo selection marker. To validate the A9 hybrids, we screened them with chromosome-specific polymorphic markers, and identified the hybrids containing either human chromosome 6, 7, 14, 18, or 21 of known parental origin. Matching paternal and maternal chromosome pairs of A9 hybrids were identified for chromosomes 6, 7, 14, and 18. The paternal-specific expression of ZAC (zinc finger protein, which regulates apoptosis and cell cycle arrest) and HYMAI (hydatidiform mole-associated and imprinted transcript), and the maternal-specific methylation of a CpG island within an imprinted domain on human chromosome 6q24, were maintained in A9 hybrids. For an example, we profiled the expression of expressed sequence tags (ESTs) and the methylation of CpG islands in the 300-kb imprinted domain around 6q24, which may be associated with cancers and transient neonatal diabetes mellitus (TNDM). Thus, the 700 A9 hybrids should be useful for various aspects of imprinting studies.

Adrenal masses: quantification of fat content with double-echo chemical shift in-phase and opposed-phase FLASH MR images for differentiation of adrenal adenomas.

PURPOSE: To quantify fat content in adrenal lesions with double-echo chemical shift magnetic resonance (MR) imaging in a phantom study and to differentiate adrenal adenomas from other adrenal masses by assessing fat content in a clinical study. MATERIALS AND METHODS: The study consisted of two parts: a phantom study and a clinical study. To explore the effect of the T1 value on in- and opposed-phase MR images of fat-containing tissues, phantom models with various proportions of fat and gadopentetate dimeglumine concentrations were implemented. Signal intensity (SI) indexes ([SI in-phase - SI opposed-phase]/SI in-phase) were calculated with double-echo fast low-angle shot (FLASH) MR imaging. In the clinical study, 23 patients with 28 adrenal masses (16 adrenal adenomas, nine adrenal metastases, and three pheochromocytomas) underwent double-echo FLASH MR imaging, and SI indexes were calculated. RESULTS: SI index reached a maximum of 0.87 at 53% fat fraction for gadopentetate dimeglumine concentration at 0.5 mmol/L as the simulated T1 of the adrenal mass. The SI indexes of the adrenal adenomas, adrenal metastases, and pheochromocytomas, respectively, were 0.36, -0.15, and -0.07, and estimated fat fraction from the phantom study was 26.5%, 0%, and 0%. All adrenal adenomas contained fat on double-echo FLASH images. There was no overlap in SI index between adenomas and other tumors. CONCLUSION: Preliminary experience indicates that quantitative measurement of the fat fraction of adrenal masses is possible with the double-echo chemical shift FLASH technique and allows for differentiating adrenal adenomas from other adrenal masses.

Fast breath-hold T2-weighted MRI of the kidney by means of half-Fourier single-shot turbo spin echo: comparison with high resolution turbo spin echo sequence.

PURPOSE: The value of the fast half-Fourier single-shot turbo spin echo (HASTE) sequence in T2-weighted MRI of the kidney was evaluated as a substitute for the conventional turbo spin echo (TSE) sequence. METHOD: Forty-five patients with suspected abnormalities of the kidney underwent MRI with a 1.5 T system. Breath-hold HASTE and respiratory-triggered TSE sequences were performed. Qualitative and quantitative analyses were performed for comparison of these sequences. RESULTS: The signal-to-noise ratio (SNR) with HASTE was higher than that with TSE. The lesion-to-kidney contrast-to-noise ratio for solid masses with HASTE was almost equal to that with TSE. For cystic masses, the CNR with HASTE was significantly higher than that with TSE (p < 0.05). Respiratory and chemical shift artifacts were significantly smaller on HASTE than on TSE (p < 0.01). However, the blurring artifact was higher on HASTE than on TSE (p = 0.01). CONCLUSION: The HASTE sequence generates high contrast images and is free of motion and chemical shift artifacts, with much better time efficacy. The sequence provides comparable diagnostic information to TSE sequences.

Effects of lysozyme and bovine serum albumin on membrane characteristics of dipalmitoylphosphatidylglycerol liposomes.

The effects of adsorption of two kinds of proteins on the membrane characteristics of liposomes were examined at pH 7.4 in terms of adsorption amounts of proteins on liposomes, penetrations of proteins into liposomal bilayer membranes, phase transition temperature, microviscosity and permeability of liposomal bilayer membranes, using positively charged lysozyme (LSZ) and negatively charged bovine serum albumin (BSA) as proteins and negatively charged L-alpha-dipalmitoylphosphatidylglycerol (DPPG) liposomes. The saturated adsorption amount of LSZ was 720 g per mol of liposomal DPPG, while that of BSA was 44 g per mol of liposomal DPPG. The penetration of LSZ into DPPG lipid membranes was greater than that of BSA. The microviscosity in the hydrophobic region of liposomal bilayer membranes increased due to adsorption (penetration) of LSZ or BSA, while the permeability of liposomal bilayer membranes increased. The gel-liquid crystalline phase transition temperature of liposomal bilayer membranes was not affected by adsorption of LSZ or BSA, while the DSC peak area (heat of phase transition) decreased with increasing adsorption amount of LSZ or BSA. It is suggested that boundary DPPG makes no contribution to the phase transition and that boundary DPPG and bulk DPPG are in the phase-separated state, thereby increasing the permeability of liposomal bilayer membranes through adsorption of LSZ or BSA. A possible schematic model for the adsorption of LSZ or BSA on DPPG liposomes was proposed.

Clinical significance of micrometastases in axillary lymph nodes assessed by reverse transcription-polymerase chain reaction in breast cancer patients.

We evaluated the clinical significance of micrometastases in axillary lymph nodes (AxLNs) of breast cancer patients for prediction of prognosis. Archived formalin-fixed paraffin-embedded AxLN specimens from 129 node-negative breast cancer patients diagnosed by routine H&E staining between 1986 and 1990 were subjected to carcinoembryonic antigen-specific reverse transcription-PCR analysis. Micrometastases were detected in 40 of 129 (31.0%) node-negative breast cancer patients. After a median follow-up period of 105.6 months, log-rank test analysis indicated that 10-year disease-free and overall survival rates by Kaplan-Meier methods were significantly better in patients without micrometastases than in patients with micrometastases [disease-free survival, 87.6% versus 66.1% (P = 0.0008); overall survival, 93.7% versus 67.8% (P = 0.0024)]. The presence of micrometastases in AxLNs was revealed by multivariate analyses to be an independent and significant predictor of clinical outcome. The hazard ratio was 3.992 (95% confidence interval, 1.293-12.323; P = 0.0161) for relapse and 4.293 (95% confidence interval, 1.043-17.675; P = 0.0436) for cancer-related death. The molecular staging of AxLNs using reverse transcription-PCR is useful for prediction of clinical outcome in early-stage breast cancer patients and can provide a powerful and sensitive complement to routine histopathological analysis.

Suppressive effects of novel ferulic acid derivatives on cellular responses induced by phorbol ester, and by combined lipopolysaccharide and interferon-gamma.

A total of 23 ferulic acid (FA) derivatives were synthesized, and investigated for their inhibitory effects on 12-O-tetradecanoylphorbol-13-acetate-induced Epstein-Barr virus (EBV) activation and superoxide (O(2)(-)) generation. Most of the derivatives showed significant EBV activation suppression or cytotoxicity at a concentration of 100 microM, with FA15 as the most potent suppressor. In both assays, FA6-FA17, bearing straight- or branched-alkyl side chains, exhibited marked suppression of O(2)(-) generation, with both FA16 and FA17 being highly active, while FA itself was virtually inactive. The activity differences seen between FA16/FA17 and FA are attributable, at least in part, to their cellular incorporating efficiencies. Further, both FA15 and FA21 attenuated the expression of inducible nitric oxide synthase and cyclooxygenase-2 proteins, while FA did not. Our results suggest that these novel FA derivatives are effective chemopreventive agents.

Insertion of bacteriophage phiFSW into the chromosome of Lactobacillus casei strain Shirota (S-1): characterization of the attachment sites and the integrase gene.

The integrase gene (int) on the genome of φFSW, which is a temperate bacteriophage of Lactobacillus casei strain Shirota (formerly denoted as S-1), and the four attachment sites on the genomes of the phage and its host were characterized by sequencing. The φFSW integrase was found to belong to the integrase family of site-specific tyrosine recombinase. The attachment sites shared a 40bp common core within which an integrative site-specific recombination occurs. The common core was flanked on one side by an additional segment of high sequence similarity. An integration plasmid, consisting of int, the phage attachment site (attP), and a selectable marker, inserted stably into the bacterial attachment site (attB) within the common core, as did the complete prophage genome at a frequency of more than 10(3)/microg of plasmid DNA. This plasmid was used as a test system for a preliminary mutational analysis of int and attP. The attB common core was located within and near the end of an open reading frame that appears to encode a homolog to glucose 6-phosphate isomerase, an enzyme of the glycolytic pathway. It is unlikely that the prophage integration inactivates this protein, since a change of only the C-terminal amino acid is predicted because of the sequence similarity between attP and attB.

Hand exercise during contrast medium delivery at thoracic helical CT: a simple method to minimize perivenous artifact.

PURPOSE: The purpose of this work was to assess the effects of hand exercise on perivenous artifact caused by undiluted venous contrast material at thoracic helical CT. METHOD: Eighty patients were prospectively and randomly assigned to thoracic helical CT with (n = 42) or without (n = 38) intermittent squeezing of a hand-sized ball during delivery of a contrast material. Two radiologists graded perivenous artifact and arterial enhancement in a blind fashion. CT attenuation values were obtained by region-of-interest measurements from arteries and veins. RESULTS: Both qualitative and quantitative analyses showed statistically significant differences in the assessment of perivenous artifact (p < 0.01). Perivenous artifact from the subclavian vein was significantly reduced in patients subjected to thoracic helical CT by using a hand exercise method. CONCLUSION: Hand exercise during contrast material delivery at thoracic helical CT minimizes perivenous artifact and improves image quality.

Detection of microsatellite alterations in nipple discharge accompanied by breast cancer.

Nipple discharge in breast cancer cases was examined loss of heterozygosity (LOH). DNA samples were extracted from both supernatant and cell pellet components of the discharge, and examined for LOH at microsatellite markers, D11S1818, D11S2000, D16S402, D16S504, D16S518, D17S520, and D17S786. At least one LOH was found in either the supernatant or cell pellet in seven out of 10 patients (70%). Five of seven samples, which were cytologically negative, were LOH positive, and only one case, which was cytologically positive, showed no LOH on the markers examined. All three samples, which were judged 'negative' by CEA measurement (<400 ng/ml), were LOH positive. This method could be a useful novel diagnostic modality for nonpalpable breast cancer with nipple discharge.