Mikihiko Kawano

A high frequency of apolipoprotein E4 isoprotein in Japanese patients with late-onset nonfamilial Alzheimer's disease

Phenotypes of apolipoprotein E (apo E) were determined by the iso-electric focusing method in 42 Japanese patients with nonfamilial late-onset Alzheimers disease (AD) and 96 age-matched controls without hyperlipidemia and/or diabetes. There was a striking difference in the distribution of apo E phenotypes between patients with AD and controls (P < 0.0001). Such a difference was mostly attributable to different frequencies of phenotypes E4/3 and E3/3. The apo E4/3 phenotype was detected in 24 (57.1%) of 42 patients with AD, more than six times oftener than in nine (9.4%) of 96 controls. In contrast, apo E3/3, which is the most common apo E phenotype in various ethnic groups, was detected in only 15 (35.7%) patients with AD. These results indicate a strong association between apo E4 isoprotein and Japanese late-onset nonfamilial AD, and that apo E4 is a possible risk factor for the development of this type of AD.

SENTINEL NODE DETECTION METHOD USING CONTRAST-ENHANCED ULTRASONOGRAPHY WITH SONAZOID IN BREAST CANCER: PRELIMINARY CLINICAL STUDY

This study aimed to evaluate the usefulness of sentinel lymph node (SLN) detection in breast cancer using contrast-enhanced ultrasonography (CEUS) with subareolar Sonazoid injection. The subjects were 20 breast cancer patients. General anesthesia was induced and 2 mL of Sonazoid was injected subareolarly. After massage of the injection site, the axillary area was observed transdermally using coded phase inversion harmonic ultrasonography with mechanical indices of 0.15 to 0.19. When contrast-enhanced lymph nodes (LNs) were seen, they were defined as CE-SLN. Two other SLN detection methods, the gamma-probe-guided and dye-guided methods, were performed together. We evaluated the SLNs detected by each method to determine if they corresponded with each other and calculated the SLN detection rate. After the SLNs were resected, pathologic examinations were done. The SLN detection rate of the CEUS-guided method, the dye-guided method and the gamma-probe-guided method were 70%, 75% and 100%, respectively. There was no statistically significant difference in these rates between the CEUS-guided and dye-guided methods (p = 0.99) but the CEUS-guided method showed a significantly lower rate than the gamma-probe-guided method (p = 0.020), and dye-guided method also showed a significantly lower rate than the gamma-probe-guided method (p = 0.047). The number of CE-SLNs was 1 or 2 (average 1.1) and each took 2 to 20 (average 5.3) min to detect. The CE-SLNs corresponded grossly with SLNs detected by the gamma-probe-guided and dye-guided methods. The pathologic results indicated no metastasis from the resected SLNs in 15 of 20 cases. However, the CEUS-guided method detected 12 cases of these 15 and CE-SLNs were detected in two of the remaining five metastasis cases. In summary, in breast cancer patients, after subareolar injection of Sonazoid, contrast-enhanced LNs were observed in real time with ultrasonography. In an initial clinical study of 20 cases, the detection rate of the CEUS-guided method was less than that of the gamma-probe-guided method. It is suggested that the CEUS-guided method using Sonazoid may, with some improvements, be a useful new modality for sentinel node identification.

Involvement of the tumor necrosis factor (TNF)/TNF receptor system in leukemic cell apoptosis induced by histone deacetylase inhibitor depsipeptide (FK228).

Inhibition of histone deacetylase (HDAC) is a novel strategy for the treatment of leukemias via restoration of aberrantly silenced genes. In this study, we conducted a detailed analysis of anti‐leukemic effects of an HDAC inhibitor (HDI), depsipeptide (FK228), using myeloid leukemia cell lines HL‐60 and K562. DNA chip analysis revealed upregulation of TNF‐α mRNA and a number of molecules involved in TNF‐signaling such as TRAF‐6, caspases‐10, and ‐7 in depsipeptide‐treated HL‐60 cells, which prompted us to examine the involvement of the TNF/TNF receptor system in the anti‐leukemic effects of the drug. Upregulation of TNF‐α was induced by depsipeptide in HL‐60 and K562 cells, which expressed type I TNF receptors (TNF‐RI). Depsipeptide activated caspases‐8 and ‐10, which in turn cleave caspases‐3 and ‐7, leading to apoptotic cell death in both cell lines. Anti‐TNF‐α neutralizing antibody and short interfering RNA (siRNA) against TNF‐RI alleviated the activation of the caspase cascade and the induction of apoptosis, indicating the presence of an autocrine loop. Finally, we demonstrated that the enhanced production of TNF‐α by depsipeptide was due to transcriptional activation of the TNF‐α gene through hyperacetylation of histones H3 and H4 in its promoter region (−208 to +35). These results suggest that autocrine production of TNF‐α plays a role in the cytotoxicity of depsipeptide against a subset of leukemias.

Comparative study of the cardio-ankle vascular index and ankle–brachial index between young Japanese and Mongolian subjects

Mongolian people have higher mortality and morbidity rates due to cardiovascular disease (CVD) than Japanese people. The cardio-ankle vascular index (CAVI) and ankle–brachial index (ABI) are both atherosclerosis-related indexes. Presently, there is no comparative information on CAVI and ABI among young subjects between Mongolian and Japanese people. A total of one hundred Mongolian (men: 39%, mean age: 20.9±2.2 years) and 115 Japanese volunteers (men: 39%, mean age: 22.0±1.8 years) were recruited from among university students. The body mass index (BMI), heart rate (HR), blood pressure (BP), CAVI, ABI, carotid intima–media thickness, blood total cholesterol (TC), glucose and C reactive protein levels were measured. The levels of BMI, HR and diastolic BP were significantly higher, and TC and glucose were significantly lower in the Mongolian subjects than in the Japanese subjects. The CAVI values (median (interquartile range): 6.5 (5.8–7.0) vs. 5.6 (5.2–6.0)) and ABI (1.11 (1.05–1.17) vs. 1.09 (1.05–1.15)) were significantly higher in the Mongolian subjects than in the Japanese subjects. The patterns of correlation between CAVI, ABI and other atherosclerotic parameters were different: in age-, gender- and BMI-adjustment correlation tests for CAVI and ABI, HR (r=−0.25 for CAVI and ABI) showed a correlation in the Mongolian subjects, and for ABI systolic BP (r=−0.28) showed a correlation in the Japanese subjects. These results suggest that Mongolian subjects may be at higher risk of CVD, even among younger individuals, than Japanese subjects.

Tamoxifen Inhibits Lipoprotein Activity: In vivo and in vitro Studies

Tamoxifen, a nonsteroidal antiestrogenic antitumor agent, has weak estrogen-like effects on lipid metabolism, however, the mechanism remains unknown. We previously reported that tamoxifen decreases the activity of lipoprotein lipase (LPL), a key enzyme in triglyceride metabolism, in patients with breast cancer. This study evaluated the effect of tamoxifen on LPL activity in vitro and in vivo. In experiment 1, total cholesterol, triglyceride, adipose tissue weight, and LPL activity of post-heparin plasma were measured in ovariectomized female rats with and without tamoxifen treatment. In experiment 2, purified very-low-density lipoprotein (VLDL) and purified LPL were incubated with and without tamoxifen or estrogen, and the triglycerides in VLDL were measured using an enzymatic method. In experiment 1, total cholesterol and adipose tissue weight decreased significantly in tamoxifen-treated rats (p < 0.001 and p < 0.01, respectively). Triglyceride measurements were not significantly different between the two groups, however, the LPL activity was lower in tamoxifen-treated rats (p < 0.005). In experiment 2, triglycerides in VLDL were significantly higher after VLDL and LPL were incubated with tamoxifen and estrogen (p < 0.005). We concluded that tamoxifen inhibits the hydrolytic activity of LPL in vivo and in vitro. This mechanism may explain the elevated serum triglyceride levels in some patients treated with tamoxifen.

A novel missense mutation of ABCA1 in transmembrane α-helix in a Japanese patient with Tangier disease

Tangier disease (TD) is a hereditary disorder characterized by the severe deficiency or absence of high-density lipoprotein cholesterol (HDL-C). TD is caused by mutations in the ATP-binding cassette transporter A1 (ABCA1) gene, most of which are located in the extracellular loops and nucleotide-binding domains. Here we describe the first case of TD carrying a missense mutation in a transmembrane alpha-helix of ABCA1. A 31-year-old Japanese woman had an extremely low level of HDL-C (1mg/dl) and yellowish tonsillar swelling, leading to the diagnosis of TD. The proband was homozygous for a point mutation of T4978C in exon 37, which results in the substitution of cysteine-1660 to arginine (C1660R) in the 8th transmembrane segment of ABCA1. Her parents, grandmother, and brother were found to be heterozygous for the same mutation. Both peripheral blood leukocytes from the patient and HEK293 cells transfected with T4978C-mutated ABCA1 normally expressed ABCA1 on the plasma membrane and had normal apolipoprotein A-I-binding ability. However, apolipoprotein A-I-mediated efflux of cholesterol and phospholipids was markedly diminished in HEK293 cells transfected with T4978C-mutated ABCA1. These results suggest that this mutant is normally translated and exists as a stable product with normal localization, yet is functionally defective. Cysteine-1660 appears to be a critical residue for cholesterol transport of ABCA1.

Coexistence of Papillary Thyroid Cancer and Hashimoto Thyroiditis in Children Report of 3 Cases

This report documents 3 pediatric papillary thyroid carcinoma cases with associated Hashimoto thyroiditis. In all 3 cases, hypoechoic nodules accompanied by multiple echogenic spots were noted on sonography of the thyroid. Hashimoto thyroiditis was suspected on the basis of positive thyroid autoantibody test results and pathologic examinations of thyroidectomy specimens, which revealed chronic thyroiditis with lymphocytic infiltration as the background of papillary thyroid carcinoma development. The potential for papillary carcinoma development warrants close follow‐up, and meticulous sonographic examinations must be performed in children with Hashimoto thyroiditis.

Effect of eicosapentaenoic acid agent on aggravated hypertriglyceridemia during pregnancy

Aggravated hypertriglyceridemia with a serum triglyceride of more than 1000 mg/dL is a risk of acute pancreatitis during pregnancy. However, there have been few reports on the administration of an eicosapentaenoic acid (EPA) agent for aggravated hypertriglyceridemia during pregnancy. A 29‐year‐old multiparous Japanese woman was transferred to our hospital at 29 + 0 weeks of gestation due to hypertriglyceridemia of 898 mg/dL. Because diet control was not enough, we decided to use an EPA agent, resulting in a reduction in triglyceride levels to 550 mg/dL. A male infant, weighing 2667 g, was born at 37 + 2 weeks transabdominally, and was complicated with respiratory distress syndrome. The final diagnosis was type III hyperlipoproteinemia with the apolipoprotein E3/2 phenotype and a broad β‐migrating lipoprotein on polyacrylamide gel electrophoresis of serum lipoproteins. In conclusion, an EPA agent may be a possible therapeutic approach for aggravated hypertriglyceridemia during pregnancy, although it may increase a risk of respiratory distress syndrome.

The usefulness of mesenterium thickness as an index of visceral fat accumulation.

Visceral fat syndrome, which includes visceral fat accumulation, glucose intolerance, hyperlipidemia, and hypertension, refers to a highly atherogenic state with a cluster of risk factors secondary to visceral fat accumulation. Although X-ray computed tomography (CT) is used commonly in diagnosis of visceral fat accumulation, it is expensive and exposes the patient to radiation. Some indices obtained using ultrasonography, which is fast and easy to use, have been reported. The ratio of the maximum thickness of preperitoneal fat (P) to the minimum thickness of subcutaneous fat (S), the abdominal wall fat index (P/S), is widely used in ultrasonographic diagnosis, however, this ratio tends to vary largely according to examiners. To establish a more reliable ultrasonographic index of visceral fat accumulation, we measured the thickness of the mesenterium, and tried to examine the relation between its thickness and maximum thickness of preperitoneal fat, minimum thickness of subcutaneous fat, and abdominal wall fat index, as well as body mass index, systolic and diastolic blood pressure, and levels of serum triglyceride, high-density lipoprotein cholesterol, total cholesterol, and blood sugar. The subjects were 131 individuals, 98 males and 33 females, averaging 47.0±7.9 years, with a mean body mass index of 24.1±2.6. The mesenterium was measured with a 3.5 or 3.75 MHz convex probe by longitudinal scanning on the left side of the umbilicus of the patient in the supine position. The thickness of the mesenterium correlated positively with maximum thickness of the preperitoneal fat, abdominal wall fat index, body mass index, and levels of serum triglyceride, total cholesterol, and blood sugar, and negatively with levels of high-density lipoprotein cholesterol. When the subjects were divided into a high-M group and a normal-M group, body mass index, triglyceride level, and blood sugar levels in the high-M group were significantly higher and level of high-density lipoprotein cholesterol was significantly lower than in the normal-M group. These results suggest that the thickness of mesenteium measured by ultrasonography may be useful in the diagnosis of accumulation of visceral fat.

Novel automated pulse immunoassay for human alpha-fetoprotein

Background To improve current alpha-fetoprotein (AFP) assays, which are expensive and time-consuming, a specific AFP reagent has been developed for practical use in our newly developed high-speed, highly sensitive pulse immunoassay (PIA) system, in which a latex immunoagglutination reaction is carried out under a high-frequency pulse voltage, leading to an enhanced immunological reaction. Methods We evaluated the assay performance (reproducibility, sensitivity, dilution linearity, interference) of the newly developed automated AFP PIA compared with the current AFP assay. Results Using pooled serum samples, the within-run reproducibility resulted in a correlation variation of 3.6–4.7%. The AFP assay detection limit was determined to be 2.5 μg/L. Linear sequential dilution was found up to nearly 700 μg/L. Even up to an AFP concentration of 1.0 g/L, the prozone phenomenon was not observed. Free and conjugated bilirubin, haemolytic haemoglobin, chyle and rheumatoid factor did not show any test interference. Using AFP-positive serum samples from 114 patients, the correlation between our PIA and a chemiluminescence immunoassay resulted in an excellent correlation coefficient of 0.994. Conclusions The performance of AFP reagents in the PIA device shows that the system has excellent speed and equal sensitivity and specificity compared with the most highly sensitive conventional method. Our PIA system thus appears ready for use in the clinical diagnosis setting.