Biomedical subjects

H Miki

Publications and source records attributed to H Miki.

At least 19 recordsLinked to original sources

PubMed2001

Sentinel lymph node biopsy in patients with papillary thyroid carcinoma.

BACKGROUND: It remains controversial whether modified radical neck dissection (MRND) for patients with papillary thyroid carcinoma improves prognosis. However, it is highly probable that the incidence of local recurrence is reduced by lymph node dissection. Sentinel lymph node (SLN) biopsy (SLNB) for patients with melanoma and breast carcinoma has been validated as an accurate method for assessing lymph node status. The objective of this study was to determine the feasibility of SLNB for the evaluation of cervical lymph node status in patients with papillary thyroid carcinoma. METHODS: After injection of methylene blue around the tumor in 22 patients with papillary thyroid carcinoma, blue-stained lymph nodes were dissected as SLNs. After the SLNB, all patients also underwent subtotal thyroidectomy and MRND. SLNs and other lymph nodes were investigated with regard to their number, distribution, size, lymph node status, and ratio of metastatic area. RESULTS: There was concordance between the SLN findings and the regional lymph node status in 19 of 21 patients (90.5%; 7 patients had both positive SLN and regional lymph node results, and 12 patients had both negative SLN and regional lymph node results). Two patients had negative SLN results but, in the end, had positive nonsentinel lymph nodes (NSLNs). The overall reliability rate of SLNB was 86.3% (19 of 22 patients). The authors experienced no complications with the use of methylene blue for the detection of SLNs. CONCLUSIONS: SLNB using methylene blue is feasible technically and is safe, and the findings correlate with cervical lymph node status. Therefore, SLNB is a good technique for estimating the status of cervical lymph nodes in patients with papillary thyroid carcinoma.

Adult↗

PubMed2001

WASP and N-WASP in human platelets differ in sensitivity to protease calpain.

Mutations of Wiskott-Aldrich syndrome protein (WASP) underlie the severe thrombocytopenia and immunodeficiency of the Wiskott-Aldrich syndrome. WASP, a specific blood cell protein, and its close homologue, the broadly distributed N-WASP, function in dynamic actin polymerization processes. Here it is demonstrated that N-WASP is expressed along with WASP, albeit at low levels, in human blood cells. The presence of approximately 160 nmol/L rapidly acting N-WASP molecules may explain the normal capacity of WASP-negative patient platelets for early agonist-induced aggregation and filopodia formation. Ex vivo experiments revealed a significant difference between WASP and N-WASP in sensitivity to calpain, the Ca++-dependent protease activated in agonist-stimulated platelets. Through the use of a series of calpain-containing broken cell systems, it is shown that WASP is cleaved in a Ca++-dependent reaction inhibitable by calpeptin and E64d and that N-WASP is not cleaved, suggesting that the cleavage of WASP by calpain functions in normal platelets as part of a Ca++-dependent switch mechanism that terminates the surface projection phase of blood cell activation processes.

Blood Platelets↗

PubMed2001

The mechanisms by which both heterozygous peroxisome proliferator-activated receptor gamma (PPARgamma) deficiency and PPARgamma agonist improve insulin resistance.

Peroxisome proliferator-activated receptor (PPAR) gamma is a ligand-activated transcription factor and a member of the nuclear hormone receptor superfamily that is thought to be the master regulator of fat storage; however, the relationship between PPARgamma and insulin sensitivity is highly controversial. We show here that supraphysiological activation of PPARgamma by PPARgamma agonist thiazolidinediones (TZD) markedly increases triglyceride (TG) content of white adipose tissue (WAT), thereby decreasing TG content of liver and muscle, leading to amelioration of insulin resistance at the expense of obesity. Moderate reduction of PPARgamma activity by heterozygous PPARgamma deficiency decreases TG content of WAT, skeletal muscle, and liver due to increased leptin expression and increase in fatty acid combustion and decrease in lipogenesis, thereby ameliorating high fat diet-induced obesity and insulin resistance. Moreover, although heterozygous PPARgamma deficiency and TZD have opposite effects on total WAT mass, heterozygous PPARgamma deficiency decreases lipogenesis in WAT, whereas TZD stimulate adipocyte differentiation and apoptosis, thereby both preventing adipocyte hypertrophy, which is associated with alleviation of insulin resistance presumably due to decreases in free fatty acids, and tumor necrosis factor alpha, and up-regulation of adiponectin, at least in part. We conclude that, although by different mechanisms, both heterozygous PPARgamma deficiency and PPARgamma agonist improve insulin resistance, which is associated with decreased TG content of muscle/liver and prevention of adipocyte hypertrophy.

Adipocytes↗

PubMed2001

Detection of peritoneal micrometastases of gastric carcinoma with green fluorescent protein and carcinoembryonic antigen promoter.

The aim of this study was to specifically visualize micrometastases in the peritoneal cavity, which cannot be detected by conventional methods, by using enhanced Green Fluorescent Protein (EGFP) containing carcinoembryonic antigen (CEA) promoter in an upstream position. In in vitro experiments, two cell lines from human gastric cancer, MKN45 and MKN1, and a cell line from human fibrosarcoma, HT1080, were transduced with pCEA-EGFP, which contains the CEA promoter region. MKN45 and MKN1, which expressed CEA mRNA, showed positive fluorescence after transduction of pCEA-EGFP, whereas HT1080 did not. In in vivo experiments, 7 days after 10(7) MKN45 had been injected into the peritoneal cavity of BALB/c nude mice, pCEA-EGFP was transduced in the peritoneal cavity using a fusogenic liposome with the envelope protein of Hemagglutinating Virus of Japan on the surface. On the peritoneum of the abdominal wall, fluorescent nodules were detected by fluorescence stereomicroscopy. These nodules had a minimal size of approximately 0.15 mm and could not be detected by conventional stereomicroscopy or macroscopy. They were histologically confirmed to be cancer cells by H&E staining. The results suggest that visualization of peritoneal micrometastasis of gastric cancer using CEA promoter and EGFP can offer a new strategy for diagnosis of micrometastasis.

Animals↗

PubMed2001

Identification of another actin-related protein (Arp) 2/3 complex binding site in neural Wiskott-Aldrich syndrome protein (N-WASP) that complements actin polymerization induced by the Arp2/3 complex activating (VCA) domain of N-WASP.

Neural Wiskott-Aldrich syndrome protein (N-WASP) is an essential regulator of actin cytoskeleton formation via its association with the actin-related protein (Arp) 2/3 complex. It is believed that the C-terminal Arp2/3 complex-activating domain (verprolin homology, cofilin homology, and acidic (VCA) or C-terminal region of WASP family proteins domain) of N-WASP is usually kept masked (autoinhibition) but is opened upon cooperative binding of upstream regulators such as Cdc42 and phosphatidylinositol 4,5-bisphosphate (PIP2). However, the mechanisms of autoinhibition and association with Arp2/3 complex are still unclear. We focused on the acidic region of N-WASP because it is thought to interact with Arp2/3 complex and may be involved in autoinhibition. Partial deletion of acidic residues from the VCA portion alone greatly reduced actin polymerization activity, demonstrating that the acidic region contributes to Arp2/3 complex-mediated actin polymerization. Surprisingly, the same partial deletion of the acidic region in full-length N-WASP led to constitutive activity comparable with the activity seen with the VCA portion. Therefore, the acidic region in full-length N-WASP plays an indispensable role in the formation of the autoinhibited structure. This mutant contains WASP-homology (WH) 1 domain with weak affinity to the Arp2/3 complex, leading to activity in the absence of part of the acidic region. Furthermore, the actin comet formed by the DeltaWH1 mutant of N-WASP was much smaller than that of wild-type N-WASP. Partial deletion of acidic residues did not affect actin comet size, indicating the importance of the WH1 domain in actin structure formation. Collectively, the acidic region of N-WASP plays an essential role in Arp2/3 complex activation as well as in the formation of the autoinhibited structure, whereas the WH1 domain complements the activation of the Arp2/3 complex achieved through the VCA portion.

Actin-Related Protein 2↗

PubMed2001

All kinesin superfamily protein, KIF, genes in mouse and human.

Intracellular transport is essential for morphogenesis and functioning of the cell. The kinesin superfamily proteins (KIFs) have been shown to transport membranous organelles and protein complexes in a microtubule- and ATP-dependent manner. More than 30 KIFs have been reported in mice. However, the nomenclature of KIFs has not been clearly established, resulting in various designations and redundant names for a single KIF. Here, we report the identification and classification of all KIFs in mouse and human genome transcripts. Previously unidentified murine KIFs were found by a PCR-based search. The identification of all KIFs was confirmed by a database search of the total human genome. As a result, there are a total of 45 KIFs. The nomenclature of all KIFs is presented. To understand the function of KIFs in intracellular transport in a single tissue, we focused on the brain. The expression of 38 KIFs was detected in brain tissue by Northern blotting or PCR using cDNA. The brain, mainly composed of highly differentiated and polarized cells such as neurons and glia, requires a highly complex intracellular transport system as indicated by the increased number of KIFs for their sophisticated functions. It is becoming increasingly clear that the cell uses a number of KIFs and tightly controls the direction, destination, and velocity of transportation of various important functional molecules, including mRNA. This report will set the foundation of KIF and intracellular transport research.

Animals↗

PubMed2001

Requirement of the basic region of N-WASP/WAVE2 for actin-based motility.

WASP family proteins activate nucleation by the Arp2/3 complex, inducing rapid actin polymerization in vitro. Although the C-terminal portion of WASP family proteins (VCA) activates nucleation by the Arp2/3 complex in pure systems, we find that this fragment lacks activity in cell extracts. Thus, polystyrene beads coated with VCA did not move in brain cytosol, while beads coated with N-WASP or WAVE2 did move. The basic clusters between the WH1 domain and the CRIB domain of N-WASP were critical for movement since beads coated with N-WASP or WAVE2 constructs missing the basic clusters (Delta basic) also did not move. Furthermore, VCA and N-WASP/WAVE2 Delta basic constructs were much less able than wild-type N-WASP and WAVE2 to induce actin polymerization in cytosol. All of the proteins, with or without the basic domain, were potent activators of nucleation by purified Arp2/3 complex.

Actins↗

PubMed2001

A novel neural Wiskott-Aldrich syndrome protein (N-WASP) binding protein, WISH, induces Arp2/3 complex activation independent of Cdc42.

We identified a novel adaptor protein that contains a Src homology (SH)3 domain, SH3 binding proline-rich sequences, and a leucine zipper-like motif and termed this protein WASP interacting SH3 protein (WISH). WISH is expressed predominantly in neural tissues and testis. It bound Ash/Grb2 through its proline-rich regions and neural Wiskott-Aldrich syndrome protein (N-WASP) through its SH3 domain. WISH strongly enhanced N-WASP-induced Arp2/3 complex activation independent of Cdc42 in vitro, resulting in rapid actin polymerization. Furthermore, coexpression of WISH and N-WASP induced marked formation of microspikes in Cos7 cells, even in the absence of stimuli. An N-WASP mutant (H208D) that cannot bind Cdc42 still induced microspike formation when coexpressed with WISH. We also examined the contribution of WISH to a rapid actin polymerization induced by brain extract in vitro. Arp2/3 complex was essential for brain extract-induced rapid actin polymerization. Addition of WISH to extracts increased actin polymerization as Cdc42 did. However, WISH unexpectedly could activate actin polymerization even in N-WASP-depleted extracts. These findings suggest that WISH activates Arp2/3 complex through N-WASP-dependent and -independent pathways without Cdc42, resulting in the rapid actin polymerization required for microspike formation.

Actin-Related Protein 2↗

PubMed2001

Ascitic fluid cytology of adenosarcoma of the ovary: a case report.

Extrauterine adenosarcoma is very rare and originates in the ovary, adnexa, or myometrium. Cytologic study of ascites is very important to determine clinical staging of malignant ovarian tumors and provide adequate therapy for recurrence. The cytomorphologic features of adenosarcoma have been only rarely described. A 77-yr-old woman visited a hospital with a complaint of lower abdominal pain for 1 mo. A tumor originating from the right adnexa in the pelvis, and involving the rectum, was found in surgery. In the ascitic fluid cytology, a few dispersed tumor cells with large cytoplasm and nuclei were oval-shaped, with nuclear invagination. The chromatin was finely granular; one or two nucleoli were conspicuous. To our knowledge, this is the fifteenth reported case of adenosarcoma of the ovary, and there have been no prior reports describing the cytological features of ascitic fluid cells in adenosarcoma of the ovary.

Adenosarcoma↗

PubMed2001

Determination of arterial input function using fuzzy clustering for quantification of cerebral blood flow with dynamic susceptibility contrast-enhanced MR imaging.

An accurate determination of the arterial input function (AIF) is necessary for quantification of cerebral blood flow (CBF) using dynamic susceptibility contrast-enhanced magnetic resonance imaging. In this study, we developed a method for obtaining the AIF automatically using fuzzy c-means (FCM) clustering. The validity of this approach was investigated with computer simulations. We found that this method can automatically extract the AIF, even under very noisy conditions, e.g., when the signal-to-noise ratio is 2. The simulation results also indicated that when using a manual drawing of a region of interest (ROI) (manual ROI method), the contamination of surrounding pixels (background) into ROI caused considerable overestimation of CBF. We applied this method to six subjects and compared it with the manual ROI method. The CBF values, calculated using the AIF obtained using the manual ROI method [CBF(manual)], were significantly higher than those obtained with FCM clustering [CBF(fuzzy)]. This may have been due to the contamination of non-arterial pixels into the manually drawn ROI, as suggested by simulation results. The ratio of CBF(manual) to CBF(fuzzy) ranged from 0.99-1.83 [1.31 +/- 0.26 (mean +/- SD)]. In conclusion, our FCM clustering method appears promising for determination of AIF because it allows automatic, rapid and accurate extraction of arterial pixels. J. Magn. Reson. Imaging 2001;13:797-806.

Adult↗

PubMed2001

Focal nodular hyperplasia coexistent with hemangioma and multiple cysts of the liver.

We report here a case of hepatic focal nodular hyperplasia (FNH) associated with hepatic hemangioma and multiple hepatic cysts in a 71-year-old man. He was admitted to our hospital because of body weight loss. Ultrasonography detected multiple cysts. and two tumors in the liver one, 3.5 cm and one, 1.6 cm. Color Doppler ultrasonography showed arterial signals within the large tumor. On dynamic computed tomography, the large tumor was a hypodense lesion which was enhanced during the arterial phase and almost isodense during the delayed phase: the small tumor was also a hypodense lesion, and was enhanced during both the arterial and delayed phases. On magnetic resonance imaging using superparamagnetic iron oxides, the large tumor had decreased signal intensity on the T2-weighted image. On hepatic arteriography, the feeding artery of the large tumor showed a spoke-wheel appearance and that of the small tumor showed a cotton-wool appearance. Ultrasonographically guided fine-needle aspiration biopsy of the large tumor revealed hepatocellular hyperplasia. Finally, we diagnosed the two hepatic tumors as FNH and hemangioma. There was no intracranial lesion. The cause of the patient's emaciation was psychogenic anorexia. To our knowledge, this is the first case report that describes the simultaneous occurrence of these three kinds of hepatic lesions. The pathogenesis of FNH still remains unclear, but this association suggests that FNH may arise because of a vascular anomaly.

Aged↗

PubMed2001

Early stage gastric MALT lymphoma with high-grade component cured by Helicobacter pylori eradication.

A 68-year-old woman was diagnosed with gastric lymphoma of the mucosa-associated lymphoid tissue (MALT) type with a high-grade component. Surgical treatment was recommended because of the presence of the high-grade component, but she refused surgery. As an alternative, she received Helicobacter pylori eradication treatment, which successfully induced regression of the lymphoma. She shows no sign of recurrence endoscopically and histologically, as of 29 months after the eradication treatment. Moreover, the B-cell monoclonality and Helicobacter pylori infection demonstrated at diagnosis has disappeared. This is one of the rare cases of gastric lymphoma of the MALT type with a high-grade component cured by Helicobacter pylori eradication alone.

2-Pyridinylmethylsulfinylbenzimidazoles↗

PubMed2001

Effects of a thiolate axial ligand on the pi-->pi* electronic states of oxoferryl porphyrins: a study of the optical and resonance Raman spectra of compounds I and II of chloroperoxidase.

Optical absorption and resonance Raman spectra have been investigated for enzymatic intermediates, compounds I and II, of chloroperoxidase (CPO) which contains a thiolate-ligated iron porphyrin. Compound I of CPO (CPO-I), an oxoferryl porphyrin pi cation radical, gave an apparently asymmetric single-peaked Soret band at 367 nm, for which band fitting analyses revealed the presence of two transition bands around 365 and 415 nm. Compound II of CPO (CPO-II), an oxoferryl neutral porphyrin, gave a split Soret spectrum with two bands (blue and red Soret bands) at 373 and 436 nm. Thus both CPO-I and CPO-II can be categorized as hyperporphyrins. The maximum extinction coefficients (epsilon(b) and epsilon(r)) and energies (Eb and Er) of the blue and red Soret bands of CPO-II were found to fall on an epsilon(b)/epsilon(r) versus Eb-Er correlation line derived from data reported for six-coordinate ferrous derivatives of cytochrome P450 and CPO. Corresponding data for CPO-I did not fall on the correlation line. Resonance enhancement of the FeIV=O stretching (vFeO) Raman band was found for CPO-I when Raman scattering was excited at wavelengths within both transition bands around 365 and 415 nm, while the vFeO Raman band was not identified for CPO-II at any of the excitation wavelengths examined here. These findings suggest that the thiolate axial ligand causes Soret band splitting of CPO-II through configuration interaction between the sulfur-->porphyrin e(g)* charge transfer and porphyrin a1u,a2u-->e(g)* transitions, while the FeO portion is important in determining the shape of the Soret band of CPO-I.

Chloride Peroxidase↗

PubMed2001

Establishment of the transformants expressing human cytochrome P450 subtypes in HepG2, and their applications on drug metabolism and toxicology.

Transformants with stable expression of a series of human cytochrome P450 (CYP) subtypes in the human hepatic cell line, HepG2, were established. These transformants are designated Hepc/1A1.4, Hepc/1A2.9, Hepc/2A6L.14, Hepc/2B6.68, Hepc/2C8.46, Hepc/2C9.1, Hepc/2C19.12, Hepc/2D6.39, Hepc/2E1.3-8 and Hepc/3A4.2-30, which stably expressed human CYP1A1, CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4, respectively. The expression of the CYP subtypes in the transformants was confirmed by both determination of enzyme activities and the reverse transcriptase polymerase chain reaction (RT-PCR) procedure. The apparent K(m) values of the expressed CYP subtypes for their specific substrates were close to those of human liver microsomes. In addition to their CYP activities, these transformants retained glucuronide- and sulfate-conjugating activities. Furthermore, the activities of CYP2C9, CYP2D6 and CYP3A4 were inhibited by their specific inhibitors. The cytotoxicity of acetaminophen (APAP), cyclophosphamide (CPA) and benz[a]anthracene (BA) were analyzed by CYP-expressing transformants. The cytotoxicity depended on the expression of CYP subtypes and increased in a dose-dependent manner. These results show the metabolic activation of APAP, CPA and BA by the specific CYP subtypes expressed in the transformants and demonstrate the usefulness of these transformants for in vitro metabolic and toxicological studies in human liver.

Acetaminophen↗

PubMed2001

WIP regulates N-WASP-mediated actin polymerization and filopodium formation.

Induction of filopodia is dependent on activation of the small GTPase Cdc42 and on neural Wiskott-Aldrich-syndrome protein (N-WASP). Here we show that WASP-interacting protein (WIP) interacts directly with N-WASP and actin. WIP retards N-WASP/Cdc42-activated actin polymerization mediated by the Arp2/3 complex, and stabilizes actin filaments. Microinjection of WIP into NIH 3T3 fibroblasts induces filopodia; this is inhibited by microinjection of anti-N-WASP antibody. Microinjection of anti-WIP antibody inhibits induction of filopodia by bradykinin, by an active Cdc42 mutant (Cdc42(V12)) and by N-WASP. Our results indicate that WIP and N-WASP may act as a functional unit in filopodium formation, which is consistent with their role in actin-tail formation in cells infected with vaccinia virus or Shigella.

3T3 Cells↗

PubMed2001

Involvement of platelet activation in experimental osteonecrosis in rabbits.

Osteonecrosis (ON) was produced experimentally in rabbits by intravenous injection of platelet activating factor (PAF) in combination with lipopolysaccharides (LPS) on two occasions separated by a week-long interval. Eleven of 15 rabbits (73%), with administration of both LPS (50 microg/kg) and PAF (10 microg/kg), exhibited microcirculatory injuries including extravasation of erythrocytes into sinusoidal spaces and formation of microthrombi in arterioles near regions of erythrocyte extravasation. Seven of 15 rabbits (47%), which received both LPS (50 microg/kg) and PAF (10 microg/kg), exhibited necrosis of trabeculae and 8 of 15 (53%) exhibited bone marrow necrosis. In addition, PAF receptor antagonist (0.3 and 3.0 mg/kg) significantly reduced the incidence of trabecular necrosis (0%) in this model. The findings of the present study suggest that platelet activation may play an important role in inducible ON, and that suppression of platelet activation may contribute prevention of ON.

Animals↗

PubMed2001

Endogenous PPAR gamma mediates anti-inflammatory activity in murine ischemia-reperfusion injury.

BACKGROUND & AIMS: Peroxisome proliferator-activated receptor gamma (PPAR gamma) is a nuclear receptor whose activation has been linked to several physiologic pathways including those related to the regulation of intestinal inflammation. We sought to determine whether PPAR gamma could function as an endogenous anti-inflammatory pathway in a murine model of intestinal ischemia-reperfusion (I/R) injury. METHODS: PPAR gamma-deficient and wild-type mice were examined for their response to I/R procedure. Treatment with a PPAR gamma-specific ligand was also performed. RESULTS: In a murine model of intestinal I/R injury, we observed more severe injury in PPAR gamma-deficient mice and protection against local and remote tissue injury in mice treated with a PPAR gamma-activating ligand, BRL-49653. Activation of PPAR gamma resulted in down-regulation of intercellular adhesion molecule 1 expression by intestinal endothelium and tissue tumor necrosis factor alpha messenger RNA levels most likely by inhibition of the NF-kappa B pathway. CONCLUSIONS: These data strongly suggest that an endogenous PPAR gamma pathway exists in tissues that may be amenable to therapeutic manipulation in I/R-related injuries.

Animals↗

PubMed2001

Expression of cyclin E and cyclin-dependent kinase 2 correlates with metastasis and prognosis in colorectal carcinoma.

The expression of cyclin E and cyclin-dependent kinases 2 (CDK2) in metastatic foci, the relationship of their expression with some clinicopathologic characteristics, and the correlation of their expression with prognosis remain unclear. To examine the roles of their expression in the progression of colorectal carcinoma, 21 normal mucosa, 9 hyperplastic polyps, 58 adenomas, 17 adenocarcinoma in adenomas, 203 primary cancers, 21 lymph node metastases, and 10 hepatic metastases were immunohistochemically stained with anti-cyclin E, anti-CDK2, and anti-Ki67 antibodies. In the carcinogenic process, both cyclin E and CDK2 expressions increased significantly. From the primary to the lymph node-metastatic foci, cyclin E protein remained unchanged, but CDK2 increased significantly. From the primary to the liver-metastatic foci, cyclin E apparently decreased, and CDK2 was reduced almost to zero. In primary carcinomas, the reduction of cyclin E was significantly associated with large tumor size, mucinous type, venous invasion, deep infiltration, lymph nodal metastasis, peritoneal metastasis, advanced stage, and poor prognosis. Decreased CDK2 was obviously correlated with large tumor size, venous invasion, deep infiltration, hepatic metastasis, advanced stage, and poor prognosis. Increased cyclin E protein was related to elevated CDK2, which was further linked to higher Ki67. Thus, CDK2 overexpression could facilitate lymph node metastasis. The overexpression of cyclin E and CDK2 may mainly promote the progression of early cancer. Anti-cyclin E or anti-CDK2 chemotherapy should be targeted to the cancers with such overexpression.

Adenocarcinoma↗