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Floral transition should be strictly regulated because it is one of the most critical developmental processes in plants. Arabidopsis terminal flower 2 (tfl2) mutants show an early-flowering phenotype that is relatively insensitive to photoperiod, as well as several other pleiotropic phenotypes. We found that the early flowering of tfl2 is caused mainly by ectopic expression of the FLOWERING LOCUS T (FT) gene, a floral pathway integrator. Molecular cloning of TFL2 showed that it encodes a protein with homology to heterochromatin protein 1 (HP1) of animals and Swi6 of fission yeast. TFL2 protein localizes in subnuclear foci and expression of the TFL2 gene complemented yeast swi6- mutants. These results suggested that TFL2 might function as an HP1 in Arabidopsis. Gene expression analyses using DNA microarrays, however, did not show an increase in the expression of heterochromatin genes in tfl2 mutants but instead showed the upregulation of the floral homeotic genes APETALA3, PISTILLATA, AGAMOUS and SEPALLATA3. The pleiotropic phenotype of the tfl2 mutant could reflect the fact that TFL2 represses the expression of multiple genes. Our results demonstrate that despite its homology to HP1, TFL2 is involved in the repression of specific euchromatin genes and not heterochromatin genes in Arabidopsis. [ABSTRACT FROM AUTHOR]

Purpose: We evaluated the usefulness of color Doppler flow imaging to compensate for the inadequate resolution of the ultrasound (US) monitoring during high-intensity focused ultrasound (HIFU) for the treatment of hepatocellular carcinoma (HCC)., Materials and Methods: US-guided HIFU ablation assisted using color Doppler flow imaging was performed in 11 patients with small HCC (<3 lesions, <3 cm in diameter). The HIFU system (Chongqing Haifu Tech) was used under US guidance. Color Doppler sonographic studies were performed using an HIFU 6150S US imaging unit system and a 2.7-MHz electronic convex probe., Results: The color Doppler images were used because of the influence of multi-reflections and the emergence of hyperecho. In 1 of the 11 patients, multi-reflections were responsible for the poor visualization of the tumor. In 10 cases, the tumor was poorly visualized because of the emergence of a hyperecho. In these cases, the ability to identify the original tumor location on the monitor by referencing the color Doppler images of the portal vein and the hepatic vein was very useful. HIFU treatments were successfully performed in all 11 patients with the assistance of color Doppler imaging., Conclusion: Color Doppler imaging is useful for the treatment of HCC using HIFU, compensating for the occasionally poor visualization provided by B-mode conventional US imaging.

The purpose of this study was to evaluate the usefulness of ultrasound-computed tomography (US-CT) 3D dual imaging for the detection of small extranodular growths of hepatocellular carcinoma (HCC). The clinical and pathological profiles of 10 patients with single nodular type HCC with extranodular growth (extranodular growth) who underwent a hepatectomy were evaluated using two-dimensional (2D) ultrasonography (US), three-dimensional (3D) US, 3D computed tomography (CT) and 3D US-CT dual images. Raw 3D data was converted to DICOM (Digital Imaging and Communication in Medicine) data using Echo to CT (Toshiba Medical Systems Corp., Tokyo, Japan), and the 3D DICOM data was directly transferred to the image analysis system (ZioM900, ZIOSOFT Inc., Tokyo, Japan). By inputting the angle number (x, y, z) of the 3D CT volume data into the ZioM900, multiplanar reconstruction (MPR) images of the 3D CT data were displayed in a manner such that they resembled the conventional US images. Eleven extranodular growths were detected pathologically in 10 cases. 2D US was capable of depicting only 2 of the 11 extranodular growths. 3D CT was capable of depicting 4 of the 11 extranodular growths. On the other hand, 3D US was capable of depicting 10 of the 11 extranodular growths, and 3D US-CT dual images, which enable the dual analysis of the CT and US planes, revealed all 11 extranodular growths. In conclusion, US-CT 3D dual imaging may be useful for the detection of small extranodular growths., (Copyright © 2011. Published by Elsevier Ireland Ltd.)

Purpose: We evaluated the efficacy of high-intensity focused ultrasound (HIFU) ablation for hepatocellular carcinoma (HCC), and a long-term study by follow-up multidetector-row computed tomography (CT) was conducted to evaluate the changes occurring in the treatment area following the HIFU ablation., Materials and Methods: HIFU ablation was carried out in 14 patients with small HCCs (≤3 lesions, ≤3cm in diameter). The HIFU system (Chongqing Haifu Tech) was used under ultrasound guidance. The evaluations were performed by follow-up CT at 1 week, 1, 3, 6 and 12 months after the HIFU ablation., Results: HIFU ablation was carried out successfully in 11 of the 14 patients. At 1 week after the HIFU, a peripheral rim enhancement was found in all cases (100%). This finding was persistent in 6 of the 11 cases (54.5%) at 1 month, and in 1 of the 11 (9%) cases at 3 months after HIFU ablation. In all cases, the rim enhancement disappeared by 6 or 12 months after the HIFU ablation. At the 12 months follow-up, a decrease in the diameter of the ablated lesions was found. The enhancement around the treated area was found to be persistent at the 12 months follow-up in the one case of recurrence of the treated site in which the safety margin was not sufficiently wide. During the follow-up period, there were 2 cases with residual of HCC tumors. We performed radiofrequency ablation (RFA) for these residual tumors after the HIFU ablation., Conclusion: To ascertain the cause of the peripheral enhancement on follow-up CT images after the HIFU ablation, in particular, to determine whether it might be caused by residual tumor or recurrence at the treated site, careful follow-up is important, especially in cases where the safety margin of the ablated area was not sufficiently wide., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

High-intensity focused ultrasound (HIFU) is a noninvasive method that can cause complete coagulation necrosis without requiring the insertion of any instruments. The hyperechoic grayscale change (hyperechoic region) is used as a sign that the treated lesion has been completely coagulated. The purpose of this study was to evaluate the first hyperechoic region during treatment using HIFU ablation according to various conditions, such as the sonication power, the depth of the tumor from the surface of the skin, and the shield rate. HIFU treatment was performed in 20 patients. The HIFU system (Chongqing Haifu Tech, Chongqing, China) was used under ultrasound guidance. Complete coagulation was achieved in 17 cases. Hyperechoic region were detected after HIFU ablation in 17 patients. The size of the hyperechoic region at a depth of >50 mm was significantly smaller than that at a depth of ≤50 mm. The number and power of the sonications for areas at a depth of >50 mm were significantly larger than those for areas at a depth of ≤50 mm. The number and power in cases with a shield rate of 31-60% were significantly larger than those in cases with a shield rate of 0-30%. When the shield rate was 0%, a hyperechoic region occurred, even when a maximum sonication power was not used. In all three cases with tumors located at a depth of greater than 70 mm and a shield rate of larger than 60%, a hyperechoic region was not seen. In conclusion, hyperechoic regions are easy to visualize in cases with tumors located at a depth of ≤50 mm or shield rates of 0-30%., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Purpose: We evaluated the safety and usefulness of high-intensity focused ultrasound (HIFU) assisted by ultrasound-computed tomography three-dimensional (US-CT 3D) dual imaging for the treatment of hepatocellular carcinoma (HCC)., Materials and Methods: HIFU ablation was performed in 13 patients with small HCC (≤3 lesions, ≤3 cm in diameter). The HIFU system (Chongqing Haifu Tech) was used under ultrasound guidance. By transferring the sagittal or axial plane of the 3D US and the CT volume data into the ZioM900, multiplanar reconstruction images were displayed in a manner resembling conventional monitor US to assist the HIFU treatment., Results: Overall, 69% (9/13) of the patients in whom good visualization using B-mode sonography could not be obtained because of the influence of multi-reflections, rib shadows, and unclear tumor margins were successfully treated under the guidance of US-CT 3D dual imaging. In 5 of the 13 patients, multi-reflections were responsible for the poor visualization. In 2 cases, the tumor was poorly visualized because of a rib shadow. In one case, the margin of the tumor was too unclear to be detected using ultrasography. The 3D US images obtained as part of the US-CT 3D dual imaging had a high resolution and were useful for examining the area of HCC invasion and for determining the extent of the ablation area. The CT images, which are not influenced by bone shadows or multi-reflections, were useful for detecting the tumors and for visualizing the presence of the intestines in the sonication zone. HIFU treatments were successfully performed in all the patients with the assistance of US-CT 3D dual imaging., Conclusion: US-CT 3D dual imaging is useful for HIFU treatment for HCC, compensating for the occasionally poor visualization provided by US monitor., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

High-intensity focused ultrasound (HIFU) is a noninvasive method that can cause complete coagulation necrosis without requiring the insertion of any instruments. The purpose of this study was to evaluate the safety and efficacy of HIFU treatment for small liver cancers without performing transcatheter arterial chemoembolization (TACE) or rib resection. HIFU ablation was performed without rib resection or the aid of TACE or percutaneous ethanol injection (PEI) in 12 patients with hepatocelullar carcinoma. The HIFU system (Chongqing Haifu Tech, Chongqing, China) was used under ultrasound guidance. All 12 patients completed the treatment without experiencing any adverse events. Complete coagulation was achieved by applying the sonications from the intercostal space when the tumor was located in the right lobe. After treatment, serum alanine aminotransferase (ALT) and serum aspartate aminotransferase (AST) levels were significantly higher than the baseline values; these levels recovered within 1 week. C-reactive protein (CRP) levels increased 1 week after treatment but decreased within 1 month. An epidural anesthetic provided sufficient pain suppression during the procedure. Edema of the subcutaneous tissue was detected in five cases, but the edema disappeared within 1 month. None of the patients developed acute hepatic failure, liver abscess or renal dysfunction. In conclusion, HIFU is effective for the treatment of patients with small liver cancer., (Copyright © 2011 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

Purpose: The study aimed to clarify whether contrast-enhanced (CE) three-dimensional (3D) ultrasonography (US) can depict characteristic hyperstain and washout patterns in minute nodules ≤10 mm of hepatocellular carcinoma (minute HCC), and whether it serves as a potent diagnostic modality., Methods: CE 3D US was carried out to depict hyperstain and washout of a solitary nodule ≤10 mm in patients with liver cirrhosis. When both patterns were depicted with success, CE computed tomography (CT) and CE magnetic resonance imaging (MRI) were also carried out as part of a comparative study, and then a histological specimen was obtained by needle biopsy. The final diagnosis was based on the histopathological findings and a follow-up on the growth of nodules., Results: All 12 minute nodules exhibiting hyperstain and washout showed evidence of HCC when examined by CE 3D US. The CE CT and CE MRI examinations could depict both of these patterns, characteristic of HCC, in most of the nodules but rather less successfully than CE 3D US., Conclusion: CE 3D US can depict hyperstain and washout patterns in minute HCC and serves as a potent modality for diagnosis.

Objective: We performed contrast-enhanced three-dimensional sonography (CE 3D US) with a perflubutane-based contrast agent to immediately evaluate the completeness of ablation of small hepatocellular carcinoma (HCC) lesions by extracorporeal high-intensity focused ultrasound (HIFU)., Subjects and Methods: Twenty-one HCC lesions were treated by a single ultrasound-guided HIFU ablation session, and CE 3D US was performed before, immediately after, and 1 week, and 1 month after HIFU, and contrast-enhanced CT (CE CT) or contrast-enhanced MRI (CE MRI) was performed before HIFU, 1 week and 1 month after HIFU, and during the follow-up period., Results: Immediately and 1 month after HIFU, 17 lesions were evaluated as adequately ablated by CE 3D US, and the other 4 lesions as residual tumors. One month after HIFU, 18 were evaluated as adequately ablated by CE CT or CE MRI, and the other 3 as residual tumors. The evaluation by CE 3D US immediately after HIFU and by CE CT or CE MRI 1 month after HIFU was concordant with 20 lesions. The kappa value for agreement between the findings of CE 3D US and other modalities by two blinded observers was 0.83. When the 1-month CE CT or CE MRI findings were used as the reference standard, the sensitivity, specificity, and accuracy of CE 3D US immediately after HIFU for the diagnosis of the adequate ablation were 100%, 75%, and 95%, respectively., Conclusion: CE 3D US appears to be a useful method for immediate evaluation of therapeutic efficacy of HIFU ablation of HCC lesions., (Copyright (c) 2009 Elsevier Ireland Ltd. All rights reserved.)

Adenine, adenosine-5'-monophosphate and adenosine-5'-triphosphate were separated by fluidic chip with PAA membranes using an aqueous mobile phase.

The possibility of using porous anodic alumina membranes as a column for normal-phase high performance liquid chromatography was evaluated using phenol and toluene with mobile phases having different solvent compositions.

Purpose: To demonstrate that liver metastases with a diameter of < or =40 mm show characteristic features on three-dimensional (3D) fusion sonographic images and that these sonographic findings can be correlated with histopathologic features of surgical specimens., Methods: Liver metastases measuring < or =40 mm were examined via contrast-enhanced 3D fusion sonography. The characteristic sonographic findings of 17 tumor nodules in 11 patients with a histopathologically confirmed diagnosis of liver metastases were investigated, and their correspondence to the pathologic features were examined in 12 resected nodules., Results: On sonograms, central vessels were visualized in all 17 tumor nodules and peripheral vessels were visualized in 12 nodules. On histopathologic examination, the portal triad vessels corresponded to the central vessels and were located at the center of the tumors. A thin layer of peritumoral hepatocytes showing various changes was also observed, and a group of tiny vessels were seen running along this layer of cells., Conclusions: Liver metastases measuring 40 mm or less in diameter were characterized by the presence of both central and peripheral vessels on contrast-enhanced 3D fusion sonography. There was good correlation between sonographic and histopathologic findings.

Dieldrin is a moth-proofing agent that was banned by the Stockholm Convention in 2001. The amount of dieldrin in wool products was measured by a microwave-assisted extraction (MAE) method. The optimal conditions were as follows: extraction solvent, acetone/n-hexane (1:1 v/v); extraction temperature, 110 degrees C; extraction time, 10 min; solvent volume, 25 mL. When six samples were used, dieldrin contents determined by GC with the proposed MAE agreed closely with those by the Japanese official method using GC with solvent extraction and cleanup by column chromatography. The proposed MAE has two merits. First, the pretreatment of the MAE needs only 4 h for 11 samples, while that using the Japanese official method needs 2 days for six samples. Second, the volume of organic solvents used for the proposed method was only about one-tenth of that used in the Japanese official method. Our proposed method seems to be easy and useful for daily (routine) tests. Dieldrin contents of 28 used wool products, which were obtained from local clothing shops and ordinary homes, were determined by GC with the proposed MAE, and six products contained dieldrin (0.310-175 ppm). The dry cleaning of the woolen yarn containing 175 ppm dieldrin did not remove a significant amount of dieldrin. Therefore, it seems likely that dieldrin is still distributed slightly but widely throughout the world.

The permeation fluxes of phenol, benzene sulfonate (BS) and benzene disulfonate (BDS) through a porous anodic alumina membrane with the perpendicularly oriented silica-surfactant nanochannel assembly membrane (NAM) were measured in water-ethanol mixture media. The permeation flux depended on solute charges and on solvent composition. As the ethanol ratio increased, the fluxes of BS and BDS increased and the flux of phenol decreased. The results of extraction/elution experiments also depended on the solute charges and the solvent composition. Chromatographic experiments in n-hexane showed that dipole and hydrophobic interactions affect the retention of solutes. Permeation of the solute across the NAM in water-ethanol mixture is likely to be determined by various factors such as dipole interaction, hydrophobic interaction, solvation, and anion-exchange efficiencies.

Objective: This study was conducted to determine whether contrast-enhanced 3-dimensional (3D) fusion ultrasonography with combined use of the plane shift and opacity control modes can serve as a useful tool for identifying the vascular characteristics of hepatic tumors in 3D perspective., Methods: Contrast-enhanced 3D fusion ultrasonography was performed in 31 patients with hepatic tumors after the intravenous injection of contrast media. The shapes, courses, and distributions of tumor vessels, displayed in 3D perspective, were examined to identify the characteristic vascular flow patterns of various tumors. Images before and after contrast media administration were compared for evaluation of the effectiveness of contrast enhancement with regard to tumor diameter for various types of tumors. The images were interpreted in a blinded manner by 3 physicians specializing in abdominal ultrasonography, and vascular flow patterns were determined on the basis of consensus., Results: The characteristic vascular flow patterns observed in hepatocellular carcinomas are reticular flow within the tumor and flow showing a ringlike distribution surrounding the tumor. Metastatic tumors show a vascular flow pattern running between adjacent tumor nodules, and hemangiomas show localized nodular flow at the tumor margins. Contrast enhancement was found to be useful for showing the vascular flow patterns in small hepatocellular carcinomas and hemangiomas., Conclusions: Contrast-enhanced 3D fusion ultrasonography makes it possible to display images combining the plane shift and opacity control modes to show tumor vessels, including minute vascular flow within hepatic tumors, in 3D perspective and to identify tumor-specific vascular flow patterns.

Purpose: To evaluate the toxicity and antitumor effect of carbon ion radiotherapy for hepatocellular carcinoma within a Phase I-II trial., Methods and Materials: Between June 1995 and February 1997, 24 patients with histopathologically proven hepatocellular carcinoma were treated to 15 fractions within 5 weeks in a step-wise dose-escalation study. The disease stage was Stage II in 10, IIIA in 6, and IVA in 8 patients. The Common Toxicity Criteria, Radiation Therapy Oncology Group/European Organization for the Research and Treatment of Cancer criteria, and Child-Pugh score were used to evaluate toxicity. The antitumor effect was evaluated by the tumor response, cumulative local control, and survival rates., Results: During a median follow-up of 71 months (range, 63-83 months), no severe adverse effects and no treatment-related deaths occurred. The Child-Pugh score did not increase by >2 points after the start of therapy. In 78% and 75% of all patients, the score did not increase by >1 point in the early and late phase, respectively. The overall tumor response rate was 71%. The local control and overall survival rate was 92% and 92%, 81% and 50%, and 81% and 25% at 1, 3, and 5 years, respectively., Conclusion: Carbon ion radiotherapy appears safe and effective for patients with hepatocellular carcinoma. Additional clinical studies using a larger subject group are required to confirm the therapeutic efficacy.

NADPH:protochlorophyllide oxidoreductase (POR) catalyzes the light-dependent reduction of protochlorophyllide. To elucidate the physiological function of three differentially regulated POR isoforms (PORA, PORB and PORC) in Arabidopsis thaliana, we isolated T-DNA tagged null mutants of porB and porC. The mature seedlings of the mutants had normal photosynthetic competencies, showing that PORB and PORC are interchangeable and functionally redundant in developed plants. In etiolated seedlings, only porB showed a reduction in the photoactive protochlorophyllide and the size of prolamellar bodies (PLBs), indicating that PORB, as well as PORA, functioned in PLB assembly and photoactive protochlorophyllide formation in etiolated seedlings. When illuminated, the etiolated porB seedling was able to green to a similar extent as the wild type, whereas the greening was significantly reduced under low light conditions. During greening, high light irradiation increased the level of PORC protein, and the greening of porC was repressed under high light conditions. The porB, but not porC, etiolated seedling was more sensitive to the far-red block of greening than the wild type, which is caused by depletion of endogenous POR proteins resulting in photo-oxidative damage. These results suggest that, at the onset of greening, PLBs are important for efficient capture of light energy for photoconversion under various light conditions, and PORC, which is induced by high light irradiation, contributes to photoprotection during greening of the etiolated seedlings.

Although sugar has been suggested to promote floral transition in many plant species, growth on high concentrations (5% [w/v]) of sucrose (Suc) significantly delayed flowering time, causing an increase in the number of leaves at the time of flowering in Arabidopsis. The effect of high concentrations of Suc seemed to be metabolic rather than osmotic. The delay of floral transition was due to extension of the late vegetative phase, which resulted in a delayed activation of LFY expression. In addition, growth on low concentrations (1% [w/v]) of Suc slightly inhibited flowering in wild-type plants. This delay resulted from effects on the early vegetative phase. This inhibition was more pronounced in tfl1, an early flowering mutant, than in the wild type. Although 1% (w/v) Suc was reported to promote floral transition of late-flowering mutants such as co, fca, and gi, floral transition in these mutants was delayed by a further increase in Suc concentration. These results suggest that sugar may affect floral transition by activating or inhibiting genes that act to control floral transition, depending on the concentration of sugars, the genetic background of the plants, and when the sugar is introduced. Growth on 1% (w/v) Suc did not restore the reduced expression levels of FT and SOC1/AGL20 in co or fca mutants. Rather, expression of FT and SOC1/AGL20 was repressed by 1% (w/v) Suc in wild-type background. The possible effects of sugar on gene expression to promote floral transition are discussed.

Objective: We assessed the role of dynamic CT in the evaluation of the efficacy of transarterial chemoembolization with iodized oil for hepatocellular carcinoma., Materials and Methods: We examined 41 hepatocellular carcinoma lesions (mean diameter, 5.0 cm) in 40 patients (mean age, 60.6 years) who underwent transarterial injection of iodized oil alone (n = 3) or emulsion of iodized oil and doxorubicin hydrochloride (n = 10) followed by gelatin sponge particles (n = 27) and subsequent hepatectomy. On dynamic CT performed within 3 weeks before oily transarterial chemoembolization and within 4 weeks before surgery, we calculated the rate of necrosis on the basis of the assumption that the portion that retained iodized oil represented necrosis. We also calculated the reduction rate of the tumor. CT findings were compared with pathologic findings of resected specimens., Results: Pathologic specimens and the necrosis rate measured on CT showed a good correlation (r = 0.83) when the portion of tumor that retained iodized oil was considered necrosis. No correlation existed if the portion that retained iodized oil was considered viable. We noted no significant correlation (r = 0.38) between the reduction rate of the tumor and necrosis rate. Also, we noted no correlation (r = 0.52) between the interval between transarterial oily chemoembolization and surgery and the reduction rate of the tumor., Conclusion: CT is suitable for the evaluation of the efficacy of oily chemoembolization for hepatocellular carcinoma on the basis of the assumption that the portion of tumor that retains iodized oil is necrotic. The rate of tumor size reduction measured on CT did not correlate with the therapeutic effect of chemoembolization.

Objective: Ideally, the diagnosis of irritable bowel syndrome (IBS) would be achieved using a minimal number of procedures. It is presumed that bowel gas is related to IBS, and it is easily visualized by plain abdominal radiograph. In the present study, to clarify the relationship between IBS and the quantity of bowel gas, the measured bowel gas volume using plain abdominal radiographs was compared with the pathology of IBS., Methods: Plain abdominal radiographs were digitized and transmitted to a computer (computed radiography) in 30 IBS patients and 30 normal controls. The quantity of bowel gas, determined as the pixel value on images and standardized by physique, was defined as the gas volume score (GVS). Using the mean +/- 2SD of GVS in the control group as the normal score, IBS patients were divided into three groups: high, normal, and low. To examine the sequential reproducibility of a similar quantity of bowel gas, a second plain abdominal radiography was performed about 2 months later, and the GVS were compared. The colonic transit time was determined using radiopaque markers., Results: There was a strong correlation between the quantities of bowel gas measured by two independent gastroenterologists. The mean GVS of IBS patients was significantly higher than that in the control group (p < 0.001). The sequential reproducibility was recognized in all 10 IBS patients. There was no significance between colonic transit time and GVS, nor between symptoms and GVS., Conclusions: Abdominal gas was analyzed objectively by using GVS, and GVS was considered to represent a useful tool for the diagnosis of IBS.

Background: Features of enhanced color flow images in small hepatocellular carcinoma (HCC) are not fully elucidated. The purpose of this study was to clarify the characteristic vascular images in small HCC observed by enhanced color Doppler., Methods: Enhanced color Doppler using the contrast agent Levovist was performed on 13 patients with HCC smaller than 30 mm. Enhanced color flow appearance was compared with angiographic findings. Time-intensity changes after injection of the contrast agent were analyzed in HCC nodules., Results: Significant improvement in the detection of color flow signals was obtained in small HCC using Levovist, from 33% in precontrast to 92% in postcontrast (p < 0.005). Three patterns of enhanced color flow images, which were related to the angiographic findings, were observed. The time-intensity curve was classified into two types by "time to peak" and "time on plateau" and was associated with the patterns of enhanced images., Conclusion: Enhanced color flow imaging promises to be a useful method for evaluating tumor vascularity noninvasively and to contribute to the elucidation of the hemodynamics in small HCC.

Two types of vacuolar sorting signals (VSSs), an asparagine-proline-isoleucine-arginine-leucine (NPIRL)-related VSS in the N-terminal propeptides (NTPPs) and a C-terminal VSS in the C-terminal propeptides (CTPPs), function differently in plant cells. A precursor to a 20-kDa protein of potato tuber (PT20) contains two NPIRL-related sequences, NPINL in a short NTPP and NPLDV close to the C terminus of the precursor. We made mutant forms of sweet potato sporamin (SPO), nPT20-SPO, in which the N-terminal pre-pro part was exchanged with that of the precursor to PT20, and SPO-PT20c, in which the C-terminal 13 amino acids of the precursor to PT20 was attached to the C terminus of delta pro-SPO which lacked NTPP. Both nPT20-SPO and SPO-PT20c were efficiently transported to the vacuoles in tobacco cells. Unlike nPT20-SPO, the vacuolar transport of SPO-PT20c was inhibited by wortmannin and by the C-terminal addition of Gly or Gly-Gly suggesting its similarity to the vacuolar transport of sporamin mediated by CTPP of barley lectin. Further analysis of the C-terminal sequence of PT20 indicated that the most C-terminal SFKQVQ sequence functions as the C-terminal VSS. These results suggest that the precursor to PT20 contains both NPIRL-like VSS in its NTPP and C-terminal VSS at the C terminus.

Thiuram (3 microM), a fungicide, was incubated in deionized water by adding 0-100 mg/L free chlorine at 30 degrees C for 30 min, and the solution was analyzed by HPLC and IC. The byproducts were identified by LC/MS, EI-MS, infrared, and (13)C NMR spectra and a reduction technique using 2-mercaptoethanol. On the basis of these results, it was found that the oxidation of thiuram with sodium hypochlorite initially produced an intermediate dimethylthiocarbamoyl dimethylcarbamoyl disulfide, which was finally degraded to bis(dimethylcarbamoyl) disulfide, its trisulfide, and dimethylamine. Subsequently, it was suggested that monitoring of bis(dimethylcarbamoyl) disulfide, its trisulfide, and dimethylamine should be included for the management and control of thiuram in tap water processed by oxidation with sodium hypochlorite.

The plasma membrane fraction from leaves of tobacco contains a 54-kDa protein with autophosphorylation activity, and the level of this protein increases after feeding of leaves with sucrose [Ohto and Nakamura (1995) Plant Physiol. 109: 973]. The 54-kDa autophosphorylation protein could not be released from the plasma membrane by treatment with salt or alkali but could be efficiently solubilized by 1% sodium deoxycholate (NaDOC). Ion-exchange chromatography of the NaDOC-solubilized proteins in the presence of octylglucoside separated the 54-kDa autophosphorylation protein into three peaks. The autophosphorylation activity of the 54-kDa protein in peaks I and II increased after feeding of leaves with sucrose. The 54-kDa protein in the peak II fraction was enriched about 125-fold starting from the microsomal membrane fraction. The 54-kDa protein in this fraction phosphorylated histone IIIS in a calcium-dependent manner and cross-reacted with an antibody against a calcium-dependent protein kinase (CDPK) of Arabidopsis thaliana. These results suggest that the 54-kDa protein in the peak II fraction is a novel isoform of CDPK which is associated with the plasma membrane and is inducible by sucrose.

The Arabidopsis LEAFY COTYLEDON1 (LEC1) gene is required for the specification of cotyledon identity and the completion of embryo maturation. We isolated the LEC1 gene and showed that it functions at an early developmental stage to maintain embryonic cell fate. The LEC1 gene encodes a transcription factor homolog, the CCAAT box-binding factor HAP3 subunit. LEC1 RNA accumulates only during seed development in embryo cell types and in endosperm tissue. Ectopic postembryonic expression of the LEC1 gene in vegetative cells induces the expression of embryo-specific genes and initiates formation of embryo-like structures. Our results suggest that LEC1 is an important regulator of embryo development that activates the transcription of genes required for both embryo morphogenesis and cellular differentiation.

Seventy-five cirrhotic patients with hyperammonemia in the past or at the time of the study were randomly divided into two groups (treated with lactulose or nontreated) in 14 hospitals in Japan. Thirty-six cirrhotic patients were diagnosed as having subclinical hepatic encephalopathy (SHE), and 39 were diagnosed as non-SHE. SHE was diagnosed when the results of all three of the quantitative psychometric tests used (number connection test, and symbol digit and block design tests of the Wechsler adult intelligence scale [revised]) were abnormal as compared with age-matched normal values. The mean number of abnormal psychometric test results and the prevalence of SHE were used for a quantitative evaluation of the efficacy of the lactulose treatment. Twenty-two of the SHE patients were treated with lactulose (45 mL/d) for 8 weeks, and the other 14 SHE patients did not receive lactulose. In the SHE patients administered lactulose, the results of the quantitative psychometric evaluation were significantly improved at 4 and 8 weeks after the beginning of the lactulose administration. The SHE had disappeared in 10 (50%) of the 20 treated patients at week 8, but it persisted in 11 (85%) of the untreated 13 patients. We concluded that lactulose treatment in cirrhotic patients with SHE is effective with respect to psychometric tests.

Objectives: To clarify the relationship between open heart surgery and gallstone formation., Methods: Fifty-one patients without gallstones (Group A) underwent cardiac surgery using a heart-lung machine and were followed for 24 months by ultrasonography. Blood tests of hemolysis markers were examined before, immediately after, and 3, 6, and 12 months after surgery. And 52 healthy candidates without gallstone (Group B) also were followed for 24 months., Results: The cumulative gallstone incidence in Group A was 15.7% at 3 months after surgery, 23.9% at 6 months, and 30.4% at and beyond 12 months and was significantly higher than that of Group B (p < 0.01). The stones showed a high dense pattern, indicative of pigment stones, in eight of the 10 patients assessed by CT. With respect to the type of surgery, latent hemolysis was seen only in patients who underwent mechanical valve replacement. However, there were no significant differences in the gallstone incidence between the patients who underwent mechanical valve replacement and those who underwent another cardiac surgery. The values of hemoglobin, haptoglobin, and lactate dehydrogenase showed abnormal values immediately after surgery, regardless of mechanical valve replacement or another cardiac surgery., Conclusions: The use of a heart-lung machine, which produces hemolysis, appears to have a close relation to gallstone formation after open cardiac surgery.

The clinical findings in 26 patients in whom hepatocellular carcinoma (HCC) was detected after the start of interferon (IFN) therapy for chronic hepatitis C were analysed. Histological study before IFN therapy showed that 34.6% of patients were categorized as stage 3 (septal fibrosis with architectural distortion; the 0-4 scale) and 80.8% demonstrated at least some evidence of septal fibrosis or more advanced features. The AFP levels examined before IFN therapy were more than 20 ng/mL in 13 patients (84.6% of those studied). One of 26 patients had a complete response to IFN therapy, while six of 26 patients had only a partial response. HCC was detected within 1 year after the start of IFN therapy in 76.9% of patients. Thus, the possibility of the early occurrence of HCC or its existence at the time of therapy should be seriously considered when IFN therapy is contemplated. Patients with stage 3 or 3-4 histology may already have a small undetectable HCC before IFN therapy. Thus, for this reason, every patient treated with IFN should be examined at short regular intervals for the development of HCC during and after IFN therapy.

Background/aims: Duck hepatitis B virus is a member of the hepadnavirus family, which possesses strong hepatotropism. Duck hepatitis B virus DNA serves as a replicative template for producing biologically active virus particles after transfection into cell lines established from human hepatocellular carcinoma or into duck liver by direct injection of calcium phosphate-precipitated DNA. Our aim was to develop a new method of liver-specific gene expression after intravenous DNA delivery., Methods/results: We inoculated duck hepatitis B virus DNA with and without cationic liposomes, Lipofectin or LipofectAMINE, as DNA carries. Two weeks after a single intravenous injection of 10 or 50 micrograms of plasmid DNA containing a head-to-tail dimer of duck hepatitis B virus DNA into 25 one-day old ducklings, duck hepatitis B virus RNA transcripts including the pregenome replicative intermediate were detected by Northern blot in the liver of eight ducks (100%) of the Lipofectin group, five ducks (63%) of the LipofectAMINE group, and three ducks (50%) of the group which received DNA without carrier. Duck hepatitis B virus RNA transcription was almost exclusively liver specific, even though the liposomes had no tissue specificity. Replicative forms of duck hepatitis B virus DNA were detected in the liver and DHBsAg was observed in the cytoplasm of the hepatocytes by immunostaining. The serum of transfected ducklings contained virus particles which were infectious in other ducklings., Conclusion: The efficient and liver-specific expression of inoculated DNA was due to the amplification of nucleic acids by active virus replication process under the control of hepatocyte specific regulation.

Background & Aims: Recent studies have shown that some variant forms of CD44, a transmembrane glycoprotein expressed on various cell surfaces, might be involved in tumor progression or tumor metastasis. The aim of this study was to analyze the expression of CD44-messenger RNA (mRNA) in colorectal cancer and colorectal adenoma to further elucidate the role of CD44 in colorectal tumorigenesis., Methods: The expression of CD44-mRNA was examined in 90 specimens from 44 patients with colorectal cancer or colorectal adenomatous polyps and in the peripheral blood leukocytes from 7 healthy volunteers by reverse transcription-polymerase chain reaction and Southern blot hybridization., Results: Strong expression of the epithelial form and variant forms containing exons 11 or 12 of the CD44 gene, which conferred metastatic behavior to rat cells, was detected in primary and metastatic tumor tissues, whereas it was very weak or not detectable in normal colonic mucosae, normal liver tissue, or peripheral blood leukocytes. However, adenomatous colorectal polyps also showed as strong an expression of epithelial and variant forms of CD44 as primary and metastatic tumor tissues., Conclusions: These results suggest that variant forms of CD44-mRNA might be expressed in an early stage of colorectal carcinogenesis.

Background/aims: A problem in pharmacotherapy for bleeding varices in portal hypertension is non-responders. The aim of this study was to elucidate the features of hemodynamic response to vasopressin in the gastroesophageal collateral vein in patients with esophageal varices., Methods: Flow velocity in the portal and the collateral left gastric vein was measured with an echo-Doppler flowmeter before and during infusion of vasopressin, 0.2 U/min, in 41 patients with cirrhosis and esophageal varices., Results: The decrease in flow velocity in the left gastric vein with vasopressin (-29 +/- 25%) was significantly smaller than that in the portal vein (-56 +/- 20%). There was no or only minimal change in flow velocity in the left gastric vein in 39% of the patients, especially in those with large-size varices. In 28 patients examined by portal catheterization, changes in flow velocity in the left gastric vein were correlated with portal pressure, and portal pressure in non-responders was significantly higher than that in responders (non-responders: 363 +/- 49, responders: 312 +/- 41 mmH2O, p < 0.05)., Conclusions: It was concluded that hepatofugal blood flow in the gastroesophageal collateral is not readily reduced by vasopressin. However, as the study was performed in a stable condition without variceal bleeding, whether these hemodynamic features will apply during acute variceal bleeding in patients who are known to have a poor hemodynamic response to vasopressin remains to be elucidated.

Effects of interferon treatment on hepatitis C virus were examined by investigating the presence of hepatitis C virus ribonucleic acid and anti-hepatitis C virus antibody in 70 patients with non-A, non-B chronic liver diseases. Twenty one patients were treated with three million units of interferon alfa 2a three times a week for 52 weeks, 24 patients were treated similarly for eight weeks, and 25 patients were given a placebo for eight weeks and served as control. Sixty six of 70 patients (94%) were positive for both hepatitis C virus RNA and second generation anti-hepatitis C virus antibody. Fourteen of 21 (67%) receiving the longterm treatment had a normalised alanine aminotransferase (ALT) activity, and in 12 of these hepatitis C virus ribonucleic acid became undetectable by the end of treatment and remained so during the three year follow up after the treatment. Anti-hepatitis C virus antibody determined by first generation assay became negative in one case at the end of the 52 week treatment, and in four cases at the end of the one year follow up. In contrast, only one of 24 (4%) who received the eight week treatment and only one of 25 (4%) who received the placebo had normalised ALT activities. Hepatitis C virus ribonucleic acid became negative in two patients undergoing short-term treatment and in none receiving the placebo. Thus, longterm interferon treatment seems effective in clearing hepatitis C virus from serum of patients with chronic liver disease.

Recent studies have revealed that a point mutation at codon 249 in the p53 gene predominates in hepatocellular carcinoma (HCC) cases from Southern Africa and China, where infection with hepatitis B virus (HBV) and contamination of aflatoxin B1 in food are risk factors for HCC. This unique mutation from G to T at the third base in codon 249 observed in human HCC cases is suggested to be linked to aflatoxin exposure. Six ducks with HCC, five of which were fed a diet containing aflatoxin B1 for 1-2 years, were analysed for the presence of point mutations at this codon of the p53 gene by polymerase chain reaction and direct nucleotide sequencing. None of the six ducks with HCC showed the change at this codon regardless of duck hepatitis B virus infection. This suggests that aflatoxin B1 itself might not be involved in the unique mutation at codon 249 in hepatocarcinogenesis, or that other factors coincident with aflatoxin may be responsible for this unique mutation.

The sugar-inducible expression of genes for sporamin and [beta]-amylase in leaf explants of sweet potato (Ipomoea batatas) and that of a [beta]-glucuronidase-fusion gene, with the promoter of the gene for [beta]-amylase in leaves of tobacco (Nicotiana tabacum), requires Ca2+ signaling (M. Ohto, K. Hayashi, M. Isobe, K. Nakamura [1995] Plant J 7: 297-307), and it was inhibited by staurosporin and K252a, inhibitors of protein kinases. Autophosphorylation activities of several potential protein kinases in leaves of tobacco were significantly higher in younger leaves than in mature leaves. However, the autophosphorylation activities of these proteins in mature leaves, especially those of the major autophosphorylatable proteins with apparent molecular masses of 56 and 54 kD, increased upon treatment of leaf discs with a 0.3 M solution of sucrose, glucose, or fructose, did not increase with sorbitol or mannitol treatments, and the increase by sucrose was inhibited by cycloheximide. Autophosphorylation of the 56- and 54-kD protein in vitro was dependent on Ca2+ and inhibited by staurosporine, K-252a, and by W-7. These results suggest that they belong to the family of calcium-dependent protein kinases. They were concentrated in the plasma membrane fraction and were released from membrane vesicles by high salt or with sodium carbonate. The possible functions of these sugar-inducible calcium-dependent protein kinases associated with the plasma membrane are discussed.

Hepatocyte related cell lines, namely human embryonic hepatocyte cell line (WRL68) and hepatoblastoma cell line (Hep G2), were tested for their ability to support hepatitis C virus (HCV) replication in vitro. The replicative intermediate of the minus strand HCV RNA was newly transcribed from the inoculated virus, and was stable in WRL68 cells over a period of 62 days. HCV RNA detected in the culture medium at 62 days after infection suggested a long period of virus secretion from this cell line. In Hep G2, transcription of the minus strand HCV RNA was also detected until 39 days after infection, but transcription and secretion of viral progeny could not be demonstrated. Although HCV infection of WRL68 was not high, this cell line might prove to be a useful tool for studying HCV replication in vitro.

The precore region of hepatitis B virus (HBV) is indispensable for secretion of e antigen protein. Therefore, the precore stop codon mutants may play an important role in the process of e antigen seroconversion. However, the presence of the mutants in hepatitis B e antigen positive carriers has not been fully studied because of difficulties in detecting the mutants in the presence of large amounts of wild-type viruses. To overcome this, a sensitive method has been developed to detect the presence of G to A stop codon mutants at codon 28 of precore region. Primers for polymerase chain reaction (PCR) were devised to introduce restriction enzyme site Sty I for wild-type viruses and Dde I for the mutants. The amplification products with these primers were digested with Sty I to exclude the products from wild-type viruses. The remaining amplicon from precore mutants were re-amplified, and the presence of precore mutant was confirmed with Dde I digestion. The presence of precore mutants was examined in 61 HBV carriers by the method combining PCR and restriction enzyme digestion. Approximately 0.1% of precore mutant DNA among 10(6) copies of wild-type virus DNA was detectable by this method. The presence of the precore mutants was detected in seven of 10 (70%) e antigen positive asymptomatic carriers, and in 29 of 36 (81%) e antigen positive patients with chronic liver diseases, and in all 15 (100%) anti-e antibody positive patients with chronic liver diseases. This study revealed that a small amount of the precore mutants was present in the majority of HBV carriers.

The experimental and clinical usefulness of a chemosensitivity test (Nuclear Damage Assay) was studied. Karyologic degenerative changes were observed as an indicator of drug sensitivity, in repeated arterial infusion chemotherapy (RAIC) using a reservoir for advanced hepatocellular carcinoma (HCC). In the experimental study, this sensitivity test was performed using five liver cell lines against 15 drugs. At the same time, the succinate dehydrogenase inhibition (SDI) test was also performed. Comparison of the results between these two tests gave a high consistency rate of 81%. Clinically, the karyologic sensitivity test was carried out in 135 patients with unresectable HCC. Drug sensitivity could be evaluated in as many as 89% of the total 135 patients. Of the patients, 43 received RAIC on an outpatient basis via a subcutaneously implanted reservoir. The objective response of RAIC on tumours of the 43 patients was evaluated as complete response, partial response, in 3 (9%) and 8 (23%) in 35 patients treated with positive drugs (positive group), and as 0 (0%) and 0 (0%) of 8 patients treated with negative drugs (negative group), respectively. As regards the prognosis, 1 year and 1.5 year survival rates were 70 and 45% in the positive group, and 42 and 0% in the negative group, respectively. As objective response in the positive group tended to be better than that in the negative group, and prognosis in the positive group was significantly better than that in the negative group, this sensitivity test appears to contribute to the improvement of therapeutic results if used to select drugs suitable for RAIC for advanced HCC.

Analysis of X-chromosome inactivation patterns in women has been used to assess the clonality of various tumors. In this report, we analyzed 27 liver tumors in women, including 18 samples obtained by the performance of ultrasonically guided thin-needle biopsies. By analysis of the heterogeneity of phosphoglycerate kinase gene, 11 of 27 (41%) cases were found to be heterozygous at the gene. Of these informative 11 cases with liver tumors, 7 cases were "large" tumors (> 25 mm in diameter) and 4 cases were "small" tumors (< 25 mm in diameter). All 7 large tumors showed monoclonal patterns by the phosphoglycerate kinase gene analysis. Of the 4 small tumors, 2 showed monoclonal, and 2 showed polyclonal patterns. The 2 with monoclonal patterns were pathologically diagnosed as hepatocellular carcinoma despite their small sizes (20 mm and 23 mm). Of the two with polyclonal patterns, the smallest one (15 mm) was diagnosed as benign adenomatous hyperplasia, and the other as hepatocellular carcinoma heavily infiltrated by lymphocytes. These data suggest that analysis of the methylation pattern of the phosphoglycerate kinase gene may be helpful on rare occasions in elucidating the nature of liver tumors but must in fact be used in conjunction with histological appearances to avoid errors secondary to inflammatory infiltrates.

We performed repeated arterial infusion chemotherapy (RAIC) in 114 advanced hepatocellular carcinoma (HCC) patients, using a subcutaneous reservoir implanted under ultrasonic guidance. In 60 patients, this was the initial therapy for the primary tumor and the other 54 patients being treated for recurrent tumor. One hundred and seventy-one patients with advanced HCC who had been treated by transcatheter arterial embolization (TAE) or single bolus arterial infusion chemotherapy before RAIC was available served as historical controls. In 97 patients, anticancer agents (4'-epidoxorubicin or acurarubicin) and Lipiodol emulsion were used, and in 17, anticancer agents alone were given. The response rates were 39.2% in the Lipiodol group and 17.6% in the non-Lipiodol group. The dose of Lipiodol and the degree of liver invasion were the most important factors influencing the response rate. The 1-, 2-, and 3-year survival rates were 55.0%, 30.9%, and 21.2%, respectively. The long-termsurvival was compared in relation to Child's classification and the presence or absence of portal vein tumor thrombosis (PVTT). In non-PVTT patients, the results of initial therapy and therapy for recurrence were similar, but recurrent Child's C patients showed a poorer prognosis. In PVTT patients, initial therapy had a better prognosis than treatment for recurrence, but initial Child's C patients had a poor long-termprognosis. During the observation period, no severe complications were encountered, but in Child's C patients, hepatic function sometimes deteriorated. Compared with the results in the 171 controls, RAIC was more useful for advanced HCC as initial therapy, and it was also beneficial for the treatment of recurrence after TAE.

Purpose: To evaluate the therapeutic effect of percutaneous ethanol injection (PEI) on small hepatocellular carcinoma (HCC) with computed tomography (CT)., Materials and Methods: Sixty-seven patients with histologically proved HCC 3 cm or less in diameter underwent PEI. The patients were regularly followed up with sonography and contrast material-enhanced CT for more than 1 year (range, 12-96 months). The CT findings were evaluated for three tumor types distinguished on the basis of their appearance relative to that of the surrounding liver parenchyma: type 1 = hyperattenuating at the early phase (n = 39), type 2 = iso- or hypoattenuating at the early phase and hypoattenuating at the late phase (n = 18), and type 3 = isoattenuating (not detected) at both the early and late phases (n = 10)., Results: After PEI, a necrotic area of HCC and the surrounding liver parenchyma was characterized as hypoattenuating at both early and late phases of contrast-enhanced CT, regardless of the type. When an HCC appeared to be completely necrotic within 3 months after PEI, this status was retained until the latest observation in all but three cases., Conclusion: Contrast-enhanced CT can correctly depict PEI-induced necrosis in HCC and is reliable for evaluating the therapeutic effect of PEI.

To study the characteristics of tumor blood flow, flow profiles from hepatocellular carcinomas (39 profiles) and normal hepatic arteries (23 profiles) were evaluated using velocity histograms obtained with Doppler ultrasound. The histograms were classified into three types: (1) high-peak, (2) flat, and (3) low-peak. Characteristically, the low-peak types and the flat types, with flows in opposing directions, were seen only in the tumor vessels. The turbulence in a phantom flow model was of the low-peak type. Spectral analysis revealed that the velocity profile of tumor blood flow was different from that of noncancerous flow and that tumor blood flow was characterized by turbulence.

Modern imaging modalities have allowed us to make reliable diagnosis of hepatocellular carcinoma (HCC). Most of HCCs are associated with liver cirrhosis in varying stages of severity and often accompanied by new occurrences of HCC after treatment. Now, newly developed nonsurgical therapeutic modalities are used extensively and achieve good results in terms of anti-tumor effects as well as post-treatment survival.

Piezoelectric extracorporeal litotripsy was performed in 128 symptomatic patients with radiolucent gall-bladder stones to assess the significance of disintegration in fragment clearance. Up to 10 repeat lithotripsy sessions were scheduled to achieve a fragment target size of < 3 mm. Fragmentation assessed by the size of the largest fragments after the last session was graded into three classes. I: sludge-like disintegration, 18%; II: < 3 mm (mean +/- s.d., 1.7 +/- 0.5 mm), 56%; and III: > or = 3 mm (3.3 +/- 0.6), 26%. All patients were initially subjected to lithotripsy alone. Bile acid dissolution therapy was started only when ultrasonography failed to show the evidence of decrease in the < 3 mm fragments during a 1 month follow up. Finally, 69 patients (54%) were treated by lithotripsy alone, and the remaining 59 received additional dissolution therapy at a mean period of 2.5 months after the initial lithotripsy. The rate of complete clearance in class I, II and III patients was 91, 42 and 10% at 6 months and 100, 68 and 49% at 18 months, respectively. Significant differences were noted between the three fragmentation grades (I vs II, III, P < 0.0001; II vs III, P < 0.02). The patients with complete clearance within 6 months were seen only in those treated by lithotripsy alone, while the majority (87%) of patients with complete clearance during the later period were seen in those treated by additional dissolution therapy. We conclude that a high degree of fragmentation appears to lead stones to an earlier period clearance, and reduce the need for dissolution therapy.