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Aims/introduction: Diabetic polyneuropathy (DPN) develops in the early stage of diabetes. However, no common diagnostic protocol has yet been established. Here, to verify that the flicker electroretinogram using a hand-held device can detect the early dysfunction of the peripheral nervous system in patients with diabetes, we investigated the correlation between the progression of DPN and neuroretinal dysfunction., Materials and Methods: In total, 184 participants with type 1 or 2 diabetes underwent a flicker electroretinogram (ERG) using a hand-held device RETeval™ and nerve conduction study. Participants were also evaluated for intima-media thickness, ankle-brachial index, toe brachial index and brachial-ankle pulse wave velocity. Parameters of the nerve conduction study were used to diagnose the severity according to Baba's classification. A multiple regression analysis was used to examine the associations of ERG parameters with the severity of DPN categorized by Baba's classification. Diagnostic properties of the device in DPN were evaluated using a receiver operating characteristic curve., Results: A multiple regression model to predict the severity of DPN was generated using ERG. In the model, moderate-to-severe DPN was effectively diagnosed (area under the receiver operating characteristic curve 0.692, sensitivity 56.5%, specificity 78.3%, positive predictive value 70.6%, negative predictive value 66.1%, positive likelihood ratio 2.60, negative likelihood ratio 0.56). In the patients without diabetic retinopathy, the implicit time and amplitude in ERG significantly correlated with the parameters of the nerve conduction study, brachial-ankle pulse wave velocity and intima-media thickness., Conclusions: Electroretinogram parameters obtained by the hand-held device successfully predict the severity of DPN. The device might be useful to evaluate DPN., (© 2020 The Authors. Journal of Diabetes Investigation published by Asian Association for the Study of Diabetes (AASD) and John Wiley & Sons Australia, Ltd.)

Predicting drug-induced liver injury is important in early stage drug discovery; however, an accurate prediction with existing hepatotoxicity evaluation tools is difficult. Conventional monolayer (2D) cultures have short viabilities and are therefore inappropriate for performing long-term toxicity tests. Conventionally used 200-μm spheroids also have toxicity detection limits. The goal of this study was to develop a humanized liver tissue capable of evaluating long-term toxicity with high sensitivity. Spheroids consisting of co-cultured cryopreserved primary human hepatocytes and human hepatic stellate cells were developed using a 3D bio-printer. The "3D bio-printed liver tissue", of ∼1 mm, was then used for long-term viability assessments (over 25 days) based on ATP, albumin, and urea levels. Hepatotoxicity evaluation was performed by analyzing the expression of genes involved in drug metabolism and transport over a 2-week drug exposure period. The 3D bio-printed liver tissue showed improved viability and enhanced gene expression of enzymes related to drug metabolism and transport, as compared to the controls. Additionally, the 3D bio-printed liver tissue demonstrated a high sensitivity for hepatotoxicity evaluation when combined with pathological evaluation and measurements for ATP production, and secretion of albumin and urea. In conclusion, the 3D bio-printed liver tissue was able to detect the toxicity of compounds that was, otherwise, undetected by 2D culture and conventionally used spheroids. These findings demonstrate a 3D bio-printed liver tissue with increased accuracy of hepatotoxicity prediction in the early stages of drug discovery, as compared to currently available methods.

The liver plays a central role in metabolism. Although many studies have described in vitro liver models for drug discovery, to date, no model has been described that can stably maintain liver function. Here, we used a unique, scaffold-free 3D bio-printing technology to construct a small portion of liver tissue that could stably maintain drug, glucose, and lipid metabolism, in addition to bile acid secretion. This bio-printed normal human liver tissue maintained expression of several kinds of hepatic drug transporters and metabolic enzymes that functioned for several weeks. The bio-printed liver tissue displayed glucose production via cAMP/protein kinase A signaling, which could be suppressed with insulin. Bile acid secretion was also observed from the printed liver tissue, and it accumulated in the culture medium over time. We observed both bile duct and sinusoid-like structures in the bio-printed liver tissue, which suggested that bile acid secretion occurred via a sinusoid-hepatocyte-bile duct route. These results demonstrated that our bio-printed liver tissue was unique, because it exerted diverse liver metabolic functions for several weeks. In future, we expect our bio-printed liver tissue to be applied to developing new models that can be used to improve preclinical predictions of long-term toxicity in humans, generate novel targets for metabolic liver disease, and evaluate biliary excretion in drug development.

Hemothorax due to rupture of metastatic hepatocellular carcinoma (HCC) is a very rare complication with high mortality because of uncontrollable hemorrhage. A 71-year-old man treated by transcatheter arterial embolization for HCC with massive bleeding from chest wall metastasis is reported. Enhanced computed tomography and selective intercostal angiogram showed a hypervascular mass in the right chest wall and extravasation of contrast agent. After successful transcatheter arterial embolization with gelatin sponge particles and metallic coils, the patient recovered from shock without major complication. To our knowledge, a successfully treated case of hemothorax due to rupture of metastatic HCC has not previously been described.

Purpose: We investigated the time interval from labeling to peak (TLP) of perfusion-signal intensity (SIs) in normal brain using a multiphase arterial spin-labeling (ASL) magnetic resonance imaging (MRI) technique as a fundamental study to assess the temporal characteristics of perfusion SIs., Materials and Methods: Twenty temporal phases of a pulsed ASL-MRI [QUASAR, quantitative signal targeting by alternating radiofrequency pulses (STAR) labeling of arterial regions] in single-slice imaging were performed in 9 volunteers. Perfusion SIs were measured and TLPs were calculated in 14 regions of interest (ROIs), 7 in each hemisphere: thalamus, lentiform nucleus, medial frontal cortex in the anterior cerebral artery (ACA) territory, temporal cortex in the middle cerebral artery (MCA) territory, medial occipital cortex in the posterior cerebral artery (PCA) territory, anterior watershed region (AWR) and posterior watershed region (PWR)., Results: Mean TLP varied with ROI (region and mean ± standard deviation in seconds): thalamus, 1.60 ± 0.11; lentiform nucleus, 1.52 ± 0.11; ACA territory, 1.53 ± 0.16; MCA territory, 1.59 ± 0.18; PCA territory, 1.68 ± 0.20; AWR, 1.79 ± 0.14; PWR, 2.00 ± 0.20. TLP in the PWR was significantly longer than those in all other regions except the AWR, and TLP in the AWR was significantly longer than those in the lentiform nucleus and the ACA territory., Conclusion: Our results revealed regional differences in the temporal characteristics of perfusion SIs on ASL-MRI.

Introduction: Shared rotating control acquisition can shorten the imaging time of territorial arterial spin labeling (tASL) by 33% compared with the normal control acquisition scheme but potentially results in an inaccurate estimate of vascular territories due to imperfect magnetization transfer compensation. Our purpose was to validate the accuracy of the shared rotating control acquisition method in evaluation of vascular territories., Methods: Twenty-four patients underwent tASL at a 3.0-T MRI with the conventional normal control acquisition method. Composite vascular territory maps, in which the blood flows from the right and left internal carotid arteries and the posterior circulation were encoded in red-green-blue, were generated as a normal averaged control-label scheme and as a simulated shared rotating control scheme. Two observers independently reported the most dominant territorial flow in 26 brain regions corresponding to the arterial segments at three post-labeling time points. Inter-reader and inter-method agreements were analyzed using κ statistics., Results: Overall inter-reader agreements were excellent for both the normal control and the shared rotating control methods (κ = 0.98, respectively). Overall inter-method agreement was also excellent (κ = 0.98), although relatively low agreement was noted in the bilateral posterior cerebral artery territories (κ = 0.79 to 0.93)., Conclusion: Our results suggested that tASL using shared rotating control acquisition can provide information on the vascular territories comparable to that obtained using the normal control acquisition while substantially shortening the imaging time.

Background and Purpose: MSDE preparation is a technique for black-blood imaging. Our purpose was to evaluate the usefulness of a 3D TSE sequence with MSDE preparation in detecting brain metastases by comparing it with conventional sequences., Materials and Methods: Postcontrast images of 227 patients who were suspected of having brain metastasis were prospectively obtained by using 3 T1-weighted 3D sequences: a gradient-echo sequence (MPRAGE), TSE-noMSDE, and TSE-MSDE. The number of visualized blood vessels and the lesion-to-normal CNR were compared among the 3 sequences. An observer test involving 9 radiologists was performed, and their diagnostic performance by using TSE-MSDE, MPRAGE, and combined TSE-MSDE and MPRAGE was compared by means of an FOM as an index of diagnostic performance derived by the JAFROC analysis, sensitivity, FP/case, and reading time., Results: TSE-MSDE resulted in significantly better vessel suppression than the other 2 methods. TSE with and without MSDE resulted in significantly higher CNRs than MPRAGE. In the observer test, significantly higher sensitivity and FOM as well as significantly shorter reading time were achieved by TSE-MSDE compared with MPRAGE, but FP/case was significantly higher with TSE-MSDE. Combined TSE-MSDE/MPRAGE resulted in significantly higher sensitivity and FOM and similar FP/case and reading time compared with MPRAGE alone., Conclusions: With blood vessel suppression and increased CNR, TSE-MSDE improves radiologists' performances in detecting brain metastases compared with MPRAGE, but it may increase FP results. Combined with MPRAGE, TSE-MSDE achieves high diagnostic performance while maintaining a low FP rate.

Purpose: the purpose of this study was to examine interhemispheric asymmetry in volume of the caudate nucleus and its age dependency., Methods: high-resolution T1-weighted brain magnetic resonance (MR) images were obtained for each subject using a 3-dimensional fast field-echo pulse sequence. The volumes of the bilateral caudate nuclei on MR images were measured using an automated method. Right-to-left comparison was made using paired t-test. Age-related change of right-to-left volume ratio (R/L ratio) was examined using Pearson's correlation coefficient., Results: fifty healthy right-handed Japanese male subjects (age 12 to 67 years, mean 39.6 years) were involved in this study. The volume of right caudate nucleus was larger than the left in 48 of 50 subjects (P < .001). R/L ratio increased with age (r= .420, P < .01)., Conclusions: our results confirmed the rightward volumetric asymmetry of caudate nucleus in right-handed individuals, and revealed that this asymmetry becomes notable with age.

Previous studies have demonstrated both functional and structural abnormalities in the frontal-striatal-thalamic circuits in obsessive-compulsive disorder (OCD). The purpose of this study was to assess volume abnormalities not only of gray matter (GM), but also of white matter (WM) in patients with OCD using voxel-based morphometry (VBM). Subjects consisted of 23 patients with OCD and 26 normal control subjects. All patients were drug-free for at least 2 weeks before the study. Three-dimensional T1-weighed MR images were obtained in all subjects. Optimized voxel-based morphometry was performed to detect structural difference between the two groups. The patients with OCD demonstrated a significant reduction of GM volume in the bilateral medial prefrontal cortex, right premotor area, right orbitofrontal cortex (OFC), right dorsolateral prefrontal cortex, and bilateral temporal and occipital regions. The OCD patients also showed a significant WM volume increase in the right anterior limb of the internal capsule, right orbitofrontal region, and a significant WM volume reduction in the left anterior cingulate gyrus. Our findings are consistent with previous studies implicating dysfunction of the frontal cortex including the OFC. The results suggested that WM volume abnormalities in the orbitofrontal region, anterior limb of the internal capsule, and anterior cingulate gyrus would imply abnormalities in the pathways of frontal-striatal circuits., (Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.)

We report a rare case of giant cell glioblastoma that was difficult to distinguish from cerebral metastasis. The MRI finding was a ring-enhancing well-circumscribed solitary brain tumor that was very similar to cerebral metastasis. Even when MRI results were considered together with the findings by magnet resonance spectroscopy and perfusion-weighted MRI, it was hard to distinguish between giant cell glioblastoma and cerebral metastasis before surgery. When we find a solitary ring-enhancing intracranial mass with little tendency of invasion, we need to consider the possibility of giant cell GBM as a differential diagnosis.

Objective: It is important to differentiate human herpesvirus 6 (HHV-6)-associated encephalopathy from herpes simplex encephalitis (HSE). Although these conditions are similar with regard to involvement of the mesial temporal lobe, HSE is sensitive to acyclovir but HHV-6 encephalopathy is not. We compared the imaging findings of the two conditions., Materials and Methods: We encountered eight cases of HHV-6 encephalopathy and nine cases of HSE. We divided an observation time into early, middle, and late periods defined as 0-2, 3-30, and more than 30 days from the onset of neurologic symptoms. Differences between HHV-6 encephalopathy and HSE on CT scans in the early period and in distribution and temporal changes in the affected regions on MR images in the three periods were analyzed., Results: At MRI in the early and middle periods, all eight patients with HHV-6 encephalopathy had exclusive involvement of the mesial temporal lobes, and all nine patients with HSE had involvement of both the mesial temporal lobes and the extratemporal regions (p < 0.01). Among patients who underwent head MRI, six of six with HHV-6 encephalopathy but none of six with HSE had resolution of high signal intensity on T2-weighted and FLAIR images (p < 0.01). Among patients who underwent head CT in the early period, none of the four with HHV-6 encephalopathy and six of the seven with HSE had abnormal findings, including parenchymal swelling, decreased attenuation of affected regions, and abnormal gyral enhancement (p < 0.05)., Conclusion: Serial MRI showed transient abnormal signal intensity in the mesial temporal lobes in patients with HHV-6 encephalopathy but persistent abnormal signal intensity in both the mesial temporal lobes and the extratemporal regions in patients with HSE. CT in the early period showed no abnormality in patients with HHV-6 encephalopathy but definite abnormal findings in patients with HSE. These differences may be useful in the differential diagnosis of the two conditions.

Continuous maturation of cerebral white matter (WM) in the postadolescent period is not fully understood. To elucidate the time course and location of possible postadolescent maturational changes in cerebral WM, we studied 60 healthy male subjects who were in their second to seventh decade using diffusion-weighted imaging. Mean diffusivity (MD) in subcortical WM was measured in 78 cortical regions in each subject's brain using an automated method. Regression analysis was used to model the age-related change in MD by either a linear or a quadratic function in each region. Age-related changes in subcortical MD were best modeled by either a linear function or a quadratic function in 27 regions including language-related regions, visual or multimodal areas in the bilateral occipital and temporal lobes, limbic areas including the bilateral parahippocampal gyri, and the bilateral postcentral and left precentral gyri. In these regions, the MD rapidly decreased until middle age and thereafter reached a plateau. Our results revealed microstructural changes in local subcortical WM and suggests a continuing maturational process in postoadolescent periods.

Background and Purpose: Cerebral hemodynamics abnormality in Alzheimer disease (AD) is not fully understood. Our aim was to determine whether regional hypoperfusion due to AD is associated with abnormalities in regional arterial blood volume (rABV) and regional arterial transit time (rATT) as measured by quantitative arterial spin-labeling (ASL) with multiple-delay time sampling., Materials and Methods: Nineteen patients with AD (9 men and 10 women; mean age, 74.5 +/- 8.6 years) and 22 cognitively healthy control subjects (11 men and 11 women; mean age, 72.8 +/- 6.8 years) were studied by using a quantitative ASL method with multiple-delay time sampling. From the ASL data, maps of regional cerebral blood flow (rCBF), rABV, and rATT were generated. A region of hypoperfusion due to AD was determined by statistical parametric mapping (SPM) analysis. Mean rCBF, rABV, and rATT values within the hypoperfused region were compared between the AD and control groups., Results: Despite the significantly lower rCBF (P = .0004) in patients with AD (27.8 +/- 7.1 mL/100 g/min) in comparison with control subjects (36.7 +/- 6.3 mL/100 g/min), no significant difference in rATT was observed between the control (0.48 +/- 0.09 seconds) and AD (0.47 +/- 0.10 seconds) groups. Mean rABV was lower in the AD group (0.22 +/- 0.10%) than in the control group (0.27 +/- 0.12%), though the difference did not reach the level of statistical significance., Conclusions: Our results revealed that regional hypoperfusion in AD is not associated with rATT prolongation, suggesting that the mechanism of hypoperfusion is distinct from that in cerebrovascular diseases.

Using a combination of atomic force, scanning and transmission electron microscopy, we found that avian erythrocytes infected with the avian malaria parasite Plasmodium gallinaceum develop approximately 60 nm wide and approximately 430 nm long furrow-like structures on the surface. Furrows begin to appear during the early trophozoite stage of the parasite's development. They remain constant in size and density during the course of parasite maturation and are uniformly distributed in random orientations over the erythrocyte surface. In addition, the density of furrows is directly proportional to the number of parasites contained within the erythrocyte. These findings suggest that parasite-induced intraerythrocytic processes are involved in modifying the surface of host erythrocytes. These processes may be analogous to those of the human malaria parasite P. falciparum, which induces knob-like protrusions that mediate the pathogenic adherence of parasitized erythrocytes to microvessels. Although P. gallinaceum-infected erythrocytes do not seem to adhere to microvessels in the host chicken, the furrows might be involved in the pathogenesis of P. gallinaceum infections by some other mechanism involving host-pathogen interactions.

To isolate high-tolerant plants against aluminum (Al), heavy metals and/or oxidative stresses as a final goal, screening of Al tolerant plants from a collection of 49 wild plants was first of all performed in this study. Andropogon virginicus L. and Miscanthus sinensis Anders showed high Al tolerant phenotypes (more than 35% values in both relative root growth and germination frequency even under 900 microM Al concentration) in our screening. Al tolerance mechanisms in these two plants were characterized and the results suggested that (1) a transport system of toxic Al ions from root to shoot, (2) a suppression of Al accumulation in root tip region and (3) a suppression of oxidative damages by an induction of anti-peroxidation enzymes, such as superoxide dismutase (SOD) and catalase, were involved in the tolerance mechanisms. Six wild plants [Andropogon, Miscanthus, Dianthus japonicus Thunb, Echinochloa crus-galli (L.) Beauv, Reynoutria japonica Houtt, and Sporobolus fertilis (Steud.) W. Clayton] were furthermore tested for their sensitivity against heavy metal stresses and oxidative stresses. The two high Al tolerant plants, Andropogon and/or Miscanthus, showed tolerance to Cr, Zn, diamide or hydrogen peroxide, suggesting common tolerance mechanisms among the tested stresses. Reynoutria showed tolerance to diamide and hydrogen peroxide, Sporobolus to Cr and Echinocholoa to Cd and Cu. Moreover, the collection of wild plants used in this study was a very useful kit to isolate tolerant plants against various abiotic stresses within a short period of time.

Background and Purpose: We investigated the relationship between tumor blood-flow measurement based on perfusion imaging by arterial spin-labeling (ASL-PI) and histopathologic findings in brain tumors., Materials and Methods: We used ASL-PI to examine 35 patients with brain tumors, including 11 gliomas, 9 meningiomas, 9 schwannomas, 1 diffuse large B-cell lymphoma, 4 hemangioblastomas, and 1 metastatic brain tumor. As an index of tumor perfusion, the relative signal intensity (SI) of each tumor (%Signal intensity) was determined as a percentage of the maximal SI within the tumor per averaged SI within normal cerebral gray matter on ASL-PI. Relative vascular attenuation (%Vessel) was determined as the total microvessel area per the entire tissue area on CD-34-immunostained histopathologic specimens. MIB1 indices of gliomas were also calculated. The differences in %Signal intensity among different histopathologic types and between high- and low-grade gliomas were compared. In addition, the correlations between %Signal intensity and %Vessel or MIB1 index were evaluated in gliomas., Results: Statistically significant differences in %Signal intensity were observed between hemangioblastomas versus gliomas (P < .005), meningiomas (P < .05), and schwannomas (P < .005). Among gliomas, %Signal intensity was significantly higher for high-grade than for low-grade tumors (P < .05). Correlation analyses revealed significant positive correlations between %Signal intensity and %Vessel in 35 patients, including all 6 histopathologic types (rs = 0.782, P < .00005) and in gliomas (rs = 0.773, P < .05). In addition, in gliomas, %Signal intensity and MIB1 index were significantly positively correlated (rs = 0.700, P < .05)., Conclusion: ASL-PI may predict histopathologic vascular densities of brain tumors and may be useful in distinguishing between high- and low-grade gliomas and in differentiating hemangioblastomas from other brain tumors.

Background: Intraarterial therapy (IAT) for acute cerebral infarction has been proven to be profitable. However, the criteria for the indications, the choice of the thrombolytic agents, and the use of adjunctive agents are controversial. We retrospectively analyzed the prognostic factors of IAT., Materials and Methods: From 1994 to 2003, 28 patients underwent IAT due to middle cerebral artery occlusion (17 women and 11 men; median age, 69 years old). We evaluated the following prognostic parameters: institution of treatment, degree of paralysis at visit, size of high-intensity area on diffusion-weighted images, dose of intraarterial urokinase administration, elapsed time from symptom onset to completion of IAT, presence of penetration of embolus by microcatheter and microguidewire, recanalization after IAT, intracranial hemorrhage (ICH) within 24 hours after IAT, and intravenous heparin administration after IAT. The outcome was evaluated at discharge and was classified into the following categories according to the modified Rankin Scale: independence (0 to 2), dependence (3 to 5), and death (6)., Results: Seven patients were judged to be independent, 16 patients were judged to be dependent, and five patients died. Patients with recanalization after IAT had a better outcome than those without (p < 0.05); patients with intracranial hemorrhage had a worse outcome than those without (p < 0.05); and patients with intravenous heparin administration after IAT had a better outcome in activities of daily living than those without (p < 0.05)., Conclusion: In addition to ICH and recanalization, our results suggested that intravenous heparin administration after IAT had a favorable effect on patient outcome.

Latex immune agglutination method with a multipurpose auto-analyser (the automated latex assay) was validated for determination of bovine lactoferrin (BLF) in various dairy products. Reproducibility-within-laboratory (intermediate precision) due to day for infant formula, UHT milk and yogurt supplemented with BLF at 50 mg/100 g for infant formula and UHT milk, and at 100 mg/100 g for yogurt were 1.62 to 3.10 mg/100 g. Reproducibility-within-laboratory due to analysis (morning, noon, and evening) for raw milk was 1.59 mg/100 g. Trueness, accuracy in determining known amounts added for BLF-supplemented dairy products was -4.7 to -2.0 mg/100 g. BLF concentration in raw milks was 20.3 to 21.8 mg/100 g. Although interference by the matrixes took place in infant formula and raw milk, BLF assays were accurately carried out by 2000-fold dilution or standard-addition method. Automated latex assay for BLF is simple, rapid, precise and accurate enough for a routine method in various dairy products containing BLF.

Although capsaicin exhibits antitumor activity, carcinogenic potential has also been reported. To clarify the mechanism for expression of potential carcinogenicity of capsaicin, we examined DNA damage induced by capsaicin in the presence of metal ion and various kinds of cytochrome P450 (CYP) using 32P-5'-end-labeled DNA fragments. Capsaicin induced Cu(II)-mediated DNA damage efficiently in the presence of CYP1A2 and partially in the presence of 2D6. CYP1A2-treated capsaicin caused double-base lesions at 5'-TG-3', 5'-GC-3' and CG of the 5'-ACG-3' sequence complementary to codon 273, a hotspot of p53 gene. DNA damage was inhibited by catalase and bathocuproine, a Cu(I) chelator, suggesting that reactive species derived from the reaction of H2O2 with Cu(I) participate in DNA damage. Formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine was significantly increased by CYP1A2-treated capsaicin in the presence of Cu(II). Therefore, we conclude that Cu(II)-mediated oxidative DNA damage by CYP-treated capsaicin seems to be relevant for the expression of its carcinogenicity.

Most of 148 newly registered leprosy patients in the past 10 years from 1995 to 2004 in Japan had received some examinations, such as histopathology test (85%), skin smear test (84%), estimation of PGL-I antibody titer (55%) or detection M. leprae with PCR method (41%). 17% of newly patients should receive wrong therapy, if they had not had aforesaid clinical examinations in addition to count skin lesions. Improvement of technical level at skin smear test should be required for accurate treatment program in Japan. In Japanese patients, the number has been decreasing year by year; epidemiological condition was different between in Okinawa prefecture and in others, about ratio of sex, kind of type and age group.

A survey of present conditions of the residents regarding aging, aftereffects and subsequent complication was conducted and its data were analyzed in the National Leprosarium Osima Seisho-en; located on an isolated island of the Seto Inland Sea. The results showed that they have become older with a various type of disease and most of them suffered from aftereffects caused leprosy, although after therapy. This survey also suggests that future problems in the management of our leprosarium will no doubt be to cope with the aging and to reducing our scale.

Many tropical and subtropical communicable diseases are prevalent in Myanmar still now. Leprosy also is not completely controlled in spite of making exertions by the Government of Myanmar and more than 10,000 new leprosy patients were detected every year. In response to the pressure of World Health Organization (WHO), the government of Myanmar declared to eliminate this disease by the end of 2003, and all vertical staff concerned with leprosy control program concentrate to reach the goal of elimination (Prevalence rate: less than 1.0 per 10,000 population). Leprosy Control and Basic Health Services Project will be carried out in the project sites for 5 years, that is, from April, 2000 to March, 2001. Project purpose that was mentioned in the PDM were to support the leprosy control programme in Myanmar through the strengthening of Basic Health Service system by conducting training activities and other diseases' control programmes such as TB and Measles, by fully utilizing the above training opportunities. The Project started to conduct the main activities from 2001 as follows, 1. BHS training 2. Training of microscopic diagnosis 3. Sewing training as one of social rehabilitations 4. Training of reconstructive surgery 5. Survey on disabilities of leprosy patients, etc.

In Japan, a cautious definition of clinical cure of leprosy has been used since 1988. This report presents a new definition of clinical cure for leprosy patients after multi-drug treatment is completed. When the patients complete the standard treatment published in 2000, they are defined as "clinically cured". The doctor in charge should inform the patient of the cure of the disease clearly. On the release from the treatment, it is important to explain necessary cares for protection against injuries and prevention from deformities. The patient should be careful about signs of relapse and reactions.

In the medaka, Oryzias latipes, sex is determined chromosomally. The sex chromosomes differ from those of mammals in that the X and Y chromosomes are highly homologous. Using backcross panels for linkage analysis, we mapped 21 sequence tagged site (STS) markers on the sex chromosomes (linkage group 1). The genetic map of the sex chromosome was established using male and female meioses. The genetic length of the sex chromosome was shorter in male than in female meioses. The region where male recombination is suppressed is the region close to the sex-determining gene y, while female recombination was suppressed in both the telomeric regions. The restriction in recombination does not occur uniformly on the sex chromosome, as the genetic map distances of the markers are not proportional in male and female recombination. Thus, this observation seems to support the hypothesis that the heterogeneous sex chromosomes were derived from suppression of recombination between autosomal chromosomes. In two of the markers, Yc-2 and Casp6, which were expressed sequence-tagged (EST) sites, polymorphisms of both X and Y chromosomes were detected. The alleles of the X and Y chromosomes were also detected in O. curvinotus, a species related to the medaka. These markers could be used for genotyping the sex chromosomes in the medaka and other species, and could be used in other studies on sex chromosomes.

Ad hoc committee of Japanese Leprosy Association recommends standard treatment protocol of leprosy in Japan, which is a modification of World Health Organization's multidrug therapy (WHO/MDT, 1997). For paucibacillary (PB) leprosy, 6 months treatment by rifampicin and dapsone (MDT/PB) is enough. However, for high bacterial load multibacillary (MB) leprosy, 12 months treatment seems insufficient. Thus, (A) For MB with bacterial index (BI) > or = 3 before treatment, 2 years treatment by rifampicin, dapsone and clofazimine (MDT/MB) is necessary. (A-1) When satisfactory decrease of BI (BI value decrease > or = 2 steps, or final BI < 3) is obtained after completion of 2 years MDT/MB, maintenance therapy by dapsone and clofazimine is recommended until BI negativity and loss of active lesions. (A-2) When BI decrease is not satisfactory (BI value decrease < 2 steps, or final BI > or = 3), MDT/MB should be continued until BI negativity and loss of active lesions. (B) For MB with BI < 3 or fresh MB (less than 6 months after the onset of the disease) with BI > or = 3, 1 year treatment by rifampicin, dapsone and clofazimine (MDT/MB) is necessary. (B-1) When BI become negative and active lesion is lost within one year, no maintenance therapy is necessary. (B-2) When BI is still positive or active lesion is remaining, additional therapy with MDT/MB for one more year is recommended. Brief summary of diagnosis, purpose of therapy, character of drugs, and prevention of deformity is also described.

The structure of [Arg8]vasopressin methylenedithioether ([AVP]CH2) has been determined in dimethylsulfoxide-d6. Two-dimensional DQF-COSY and NOESY spectra were measured and used to derive angle and distance constraints for restrained molecular dynamics (MD) calculations. In the MD trajectory, two types of beta-turn structure were found in the region from Tyr2 to Asn5, suggesting an equilibrium between type-I and type-II' beta-turn structures. When Halpha chemical shifts were used as an additional constraint, the type-I turn was favoured. To validate this result, an independent energy minimization procedure was used, using differences between calculated and observed chemical shifts. The two approaches gave essentially identical results. It is therefore concluded that the type-I turn predominates in solution. Analysis of calculated chemical shift contributions suggests that the beta-turn structure found in AVP is well preserved in [AVP]CH2, although the pressin ring size is expanded.

We used the combination of an atomic force microscope and a light microscope equipped with epifluorescence to serially image Plasmodium falciparum-infected erythrocytes. This procedure allowed us to determine unambiguously the presence and developmental stage of the malaria parasite as well as the number and size of knobs in singly, doubly, and triply infected erythrocytes. Knobs are not present during the ring stage of a malaria infection but a lesion resulting from invasion by a merozoite is clearly visible on the erythrocyte surface. This lesion is visible into the late trophozoite stage of infection. Knobs begin to form during the early trophozoite stage of infection and have a single-unit structure. Our data suggest the possibility that a two-unit structure of knobs, which was reported by Aikawa et al. (1996, Exp. Parasitol. 84, 339-343) using atomic force microscopy, appears to be a double-tipped image. The number of knobs per unit of host cell surface area is directly proportional to parasite number in both early and late trophozoite stages. These results indicate that knob formation by one parasite does not influence knob formation by other parasites in a multiply infected erythrocyte. In addition, knob volume is not influenced by either parasite stage or number at the late trophozoite stage, indicating that the number of component molecules per knob is constant throughout the parasite maturation process.

Both living and fixed specimens of the medically-important parasitic protozoa, Trypanosoma cruzi, Toxoplasma gondii, Giardia lamblia, Entamoeba histolytica, and Acanthamoeba spp. were studied by atomic force microscopy (AFM). The preparation of fixed specimens was similar to methods used for either scanning or transmission electron microscopy. AFM scanning was performed using both contact and tapping modes. A classical fixation procedure utilizing glutaraldehyde followed by ethanol dehydration was not suitable for all parasite species. AFM images could not be obtained from fixed samples of T. cruzi, T. gondii or E. histolytica. However, excellent topographic images could be obtained from specimens of G. lamblia and Acanthamoeba under identical conditions. Critical point drying permitted AFM imaging of both trypomastigote and epimastigote stages of T. cruzi. Phase imaging of T. cruzi elucidated unique surface details at a level of resolution not visible using any other imaging modalities. AFM elasticity map imaging of T. cruzi-infected and T. gondii-infected cells demonstrated that both parasites were markedly firmer than the surrounding host cell cytoplasm. The parasitophorous vacuole surrounding replicating T. gondii tachyzoites was also visualized by elasticity map imaging. These data suggest that although much remains to be learned about preparing parasitic protozoa for AFM imaging, the technique has the potential of providing unique and important insights into these disease causing organisms.

Atomic force microscope-based phase imaging in air is capable of elucidating variations in material properties such as adhesion, friction, and viscoelasticity. However, the interpretation of phase images of specimens in a fluid environment requires clarification. In this report, we systematically analyzed atomic force microscope-derived phase images of mica, glass, and collagen under the same conditions as used for living cells at various tapping forces; the resulting data provide critical information for the interpretation of phase images of living cells. The peripheral regions of COS-1 cells consistently show a more negative phase shift than the glass substrate in phase images at set-point amplitude: free amplitude (Asp/A0) = 0.6-0.8. In addition, at all Asp/A0 values suitable for phase imaging, tapping frequency appears to be high enough to ensure that phase shifts are governed primarily by stiffness. Consequently, phase imaging is capable of high resolution studies of the cellular surface by detecting localized variations in stiffness. We demonstrate that phase imaging of a bifurcating fiber in COS-1 cell cytoplasm is readily capable of a lateral resolution of approximately 30 nm.

The atomic force microscope (AFM) is becoming an important tool for qualitative and quantitative analyses of biological material. However, the difficulties involved in maintaining long-term, steady-state physiologic conditions and the problems associated with analyzing force curves generated from highly viscoelastic biological structures impede the use of the AFM for studies of kinetic processes in living vertebrate cells. In this report, we describe a simple method to track reproducibly kinetic changes in the localized stiffness of vertebrate cells. We tested our method on a study of vertebrate cells in mitosis and found a marked but transient decrease in stiffness occurs in the mitotic spindle region during anaphase. We propose that physical-chemical changes in the mitotic apparatus, most probably, changes in the state of polymerization of interzonal spindle fibers which also have been reported to undergo a marked reduction in birefringence during anaphase, are responsible for the observed decrease in stiffness. Our methodology affords a new approach to studying mitotic events and should be applicable to studies of a variety of viscoelastic properties of living cells.

The Meiji government that modernized Japan decided the direction which adopted western medicine positively and eliminated Oriental medicine, and carried out the policy to inhibit acupuncture (1874). However, there was no concrete system until 1885, when the Ministry of Home Affairs issued a report to entrust the permission and superintendance of acupuncture to each prefecture. On the other hand, modern education for the blind was begun in various parts of the country in 1878, and acupuncture was adopted as a vocational course. The Rakuzenkai Blind School began Anma and acupuncture in 1881, but in 1885 acupuncture was removed from the curriculum and put under the direct control of the Ministry of Education. The principal of the school, Ryokichi Yatabe, had doubts about this and sent a question to Tokyo Imperial University. The answer to it (1887) was to recognize acupuncture, and this recognition became an admission of the revival of acupuncture in the course of blind education and the basis of the thought in legislation for modern Japanese acupuncture. There is a high possibilty that the reply to Yatabe was influenced by the article of Sansaku Okumura (1864-1912), a blind man in Kanazawa City, which appeared in the 157th issue of "Iji Shimbun" (1885).

Structural and functional analysis were performed on biliverdin-associated cyanoprotein (CP) from the hemolymph (CP-1 to CP-4) and eggs (CPegg, identical to CP-1) of the bean bug, Riptortus clavatus. Isoelectric focusing analysis of purified CPegg and CP-4 revealed that they are composed of a single alpha subunit and a more acidic beta subunit respectively. N-terminal amino acid sequencing revealed that there are six different amino acid residues between the two subunits up to the 29 cycles determined, indicating that they are encoded by different genes. Appreciable sequence similarities in N-terminal region are found between the CP subunits and several insect hexamers. By chemical cross-linking analysis these CPs were demonstrated to have hexameric structures. Two dimensional peptide mapping revealed that alpha and beta subunits share common structures in part and that CP-2 and CP-3 are hybrid molecules bearing both alpha and beta subunits. From these results the molecular structure of CPs was established as follows: CP-1 (CPegg) = alpha 6; CP-2 = alpha 4 beta 2; CP-3 = alpha 2 beta 4; CP-4 = beta 6. In addition, using 125I-labeled CPegg and CP-4, in vivo incorporation into several tissues was examined during the nymphal-adult development. During the late phase of the 5th instar, CP-4 was sequestered preferentially over CPegg by the fat body, while after emergence both proteins were suggested to be incorporated into the newly-formed cuticle. CPegg was sequestered massively by the developing ovaries whereas no appreciable incorporation of CP-4 was observed.

A technique for culturing skin was devised whereby hair follicles in a normal state were generated from a single-cell suspension of embryonic rat skin. Dissociated cells obtained by trypsinization of the day-15 embryonic lip were cultured by a two-step procedure in vitro. Reorganization of hair-follicle rudiments was accompanied by reaggregation of the cells during a 24-hour initial culture with rotation, and the rudiments differentiated into hair follicles within a week during subsequent subculture of the cell aggregates by floatation. The light-microscopic features and the size of the follicles were similar to those of day-18 vibrissa follicles during normal development in vivo. Furthermore, the stratification of cells, including subcellular differentiation, and the ultrastructure of the hair follicles generated in vitro were similar to those of normal hair follicles with well-keratinized hair shafts. The present system appears to be a useful model for analytical studies in vitro on the formation of hair follicles and for studies designed to facilitate the transplantation of human hair.