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Biomedical subjects

H Koyama

Publications and source records attributed to H Koyama.

At least 37 records · Page 2Linked to original sources
PubMed2002

Relative contributions of myeloperoxidase and NADPH-oxidase to the early host defense against pulmonary infections with Candida albicans and Aspergillus fumigatus.

Generation of oxidative products by phagocytic cells is known to be an important host defense mechanism directed toward killing of invading microorganisms. The importance of two major oxidant-producing enzymes, myeloperoxidase (MPO) and NADPH-oxidase, in in vivo fungicidal action was directly compared in genetically engineered mice. Both MPO-deficient (MPO-/-) and NADPH-oxidase-deficient (X-linked chronic granulomatous disease [X-CGD]) mice showed increased susceptibility to pulmonary infections with Candida albicans and Aspergillus fumigatus compared with normal mice, and the X-CGD mice exhibited shorter survivals than MPO-/- mice. This increased mortality of X-CGD mice was associated with a 10- to 100-fold increased outgrowth of the fungi in their organs during the first 6 days. These results suggest that superoxide (O2-) produced by NADPH-oxidase is more important than hypochlorous acid (HOCl) produced by MPO, although both oxidative products obviously contribute to the host defense against pulmonary infection with those fungi. We also observed that MPO-/-/X-CGD double knockout mice showed comparable levels of susceptibility to the X-CGD mice against C. albicans and A. funigatus, indicating that MPO is unable to play a role in host defense in the absence of NADPH-oxidase. This strongly suggests that hydrogen peroxide, the precursor of HOCl, is solely derived from O2- produced by NADPH-oxidase.

Animals↗
PubMed2001

DNA ligase IV-deficient cells are more resistant to ionizing radiation in the absence of Ku70: Implications for DNA double-strand break repair.

Vertebrate cells have evolved two major pathways for repairing DNA double-strand breaks (DSBs), homologous recombination (HR) and nonhomologous DNA end-joining (NHEJ). To investigate the role of DNA ligase IV (Lig4) in DSB repair, we knocked out the Lig4 gene (LIG4) in the DT40 chicken B-lymphocyte cell line. The LIG4(-/-) cells showed a marked sensitivity to X-rays, bleomycin, and VP-16 and were more x-ray-sensitive in G(1) than late S or G(2)/M, suggesting a critical role of Lig4 in DSB repair by NHEJ. In support of this notion, HR was not impaired in LIG4(-/-) cells. LIG4(-/-) cells were more x-ray-sensitive when compared with KU70(-/-) DT40 cells, particularly at high doses. Strikingly, however, the x-ray sensitivity of KU70(-/-)/LIG4(-/-) double-mutant cells was essentially the same as that of KU70(-/-) cells, showing that Lig4 deficiency has no effect in the absence of Ku. These results indicate that Lig4 is exclusively required for the Ku-dependent NHEJ pathway of DSB repair and that other DNA ligases (I and III) do not substitute for this function. Our data may explain the observed severe phenotype of Lig4-deficient mice as compared with Ku-deficient mice.

Animals↗
PubMed2001

DNA polymerase beta is not essential for the formation of palindromic (P) region of T cell receptor gene.

Formation of palindromic (P) region at the variable (V)-diversity (D)-joining (J) junction in DNA polymerase beta (pol-beta) deficient mice were investigated by sequencing of reverse transcriptase-polymerase chain reaction (RT-PCR) products of mRNAs encoding the beta chain of T cell receptor (TCR). Total 42 and 43 cDNA clones encoding V(beta8)-D(beta)-J(beta)-C(beta) from E18.5 embryonic thymocytes of pol-beta gene knocked-out and wild type control mouse, respectively, were sequenced. Among them five and six clones from pol-beta knocked-out and wild type, respectively, have P insertions of two nucleotides. This result unequivocally indicates that pol-beta, which is one of the repair-type DNA polymerases most abundantly expressed in thymus and spleen, is not essential for the formation of P region.

Animals↗
PubMed2001

Decreased topoisomerase IIalpha expression confers increased resistance to ICRF-193 as well as VP-16 in mouse embryonic stem cells.

To elucidate the relationship between topoisomerase (topo) II expression and sensitivity to anti-topo II drugs in mammalian cells, we generated mouse embryonic stem cell mutants heterozygous for the topo IIalpha gene by gene targeting. The level of topo IIalpha in the heterozygous cells reduced to one-half of that found in wild-type cells, while topo IIbeta levels were similar in both cell types. Importantly, the heterozygous cells exhibited an increased resistance to ICRF-193 as well as VP-16, suggesting that ICRF-193, like VP-16, exerts its cytotoxicity through converting topo II to a poison.

Animals↗
PubMed2001

Fibrillar collagen specifically regulates human vascular smooth muscle cell genes involved in cellular responses and the pericellular matrix environment.

Proliferation and alpha(v)beta(3) integrin-dependent migration of vascular smooth muscle cells are suppressed on polymerized type I collagen. To identify genes specifically regulated in human smooth muscle cells by polymerized collagen, we used the suppressive subtraction hybridization technique. Compared with smooth muscle cells cultured on monomer collagen, polymerized collagen suppresses the following: (1) a number of other extracellular matrix proteins, including fibronectin, thrombospondin-1, tenascin-C, and cysteine-rich protein 61; (2) actin binding proteins including alpha-actinin; (3) signaling molecules; (4) protein synthesis-associated proteins; and (5) genes with unknown functions. Some of the identified genes, including cysteine-rich protein 61, show unique kinetics of mRNA regulation by monomer or polymerized collagen distinct from growth factors, suggesting extracellular matrix-specific gene modulation. Moreover, in vivo balloon catheter-mediated injury to the rat carotid artery induces many of the genes that are suppressed by polymerized collagen. Protein levels of thrombospondin-1 and fibronectin are also suppressed by polymerized collagen. Thrombospondin-1-mediated smooth muscle cell migration on vitronectin is significantly inhibited after culture on polymerized collagen for 24 hours, which is associated with decreased alpha-actinin accumulation at focal adhesions. Thus, polymerized type I collagen dynamically regulates gene expression, pericellular accumulation of extracellular matrix molecules, and the response to a given matrix molecule.

Actinin↗
PubMed2001

Combination technique is superior to dye alone in identification of the sentinel node in breast cancer patients.

BACKGROUND: The purpose of the present study was to evaluate whether the combination of dye and radioisotope would improve the detection rate of sentinel nodes (SN) and the diagnostic accuracy of axillary lymph node status over dye alone in patients with breast cancer. METHODS: Sentinel node biopsy (SNB) was performed in stages I or II breast cancer patients with clinically negative nodes using dye alone (indocyanine green) or a combination of dye and radioisotope (99mTc-radiolabelled tin colloid). RESULTS: SNB guided by dye alone was performed in 93 patients and SNB guided by a combination of dye and radioisotope was performed in 138 patients. The detection rate of SN was significantly (P = 0.006) higher in the combination group (94.9%) than in the dye alone group (83.9%). The sensitivity, specificity, and overall accuracy of SNB in the diagnosis of axillary lymph node status were 100, 100, and 100%, respectively, for the combination group, and 81.0, 100, and 94.9%, respectively, for the dye alone group. There were no false negatives in the combination group, but four false negatives (19.0%) in the dye alone group. The combination method was significantly superior to the dye alone method for sensitivity (P = 0.011) and accuracy (P = 0.018). CONCLUSIONS: The addition of a radioisotope to the dye in SNB increases the detection rate of SNs in breast cancer patients, and SNs detected by the combination method predict the axillary lymph node status with greater accuracy than those detected by the dye alone method.

Breast Neoplasms↗
PubMed2001

Molecular pathways of cyclic nucleotide-induced inhibition of arterial smooth muscle cell proliferation.

Cyclic adenosine 3',5'-monophosphate (cAMP) and cyclic guanosine 3',5'-monophosphate (cGMP) are second messengers involved in the intracellular signal transduction of a wide variety of extracellular stimuli. These signals regulate many biological processes including cell proliferation, differentiation, migration, and apoptosis. Recently, significant progress has been achieved in the molecular basis underlying cyclic nucleotide regulation of cell proliferation. This review summarizes our knowledge of the signaling pathways regulated by cyclic nucleotides in arterial smooth muscle cells.

Animals↗
PubMed2001

Bone resorption associated with uncoupling of osteoclastic and osteoblastic activities in adult T cell leukemia with hypercalcemia: case report.

A 64-year-old woman with adult T cell leukemia (ATL) was admitted to our hospital with severe hypercalcemia. The serum calcium level was elevated to 14.9 mg/dl. Biochemical parameters for bone formation including serum osteocalcin (bone Gla protein, BGP) and alkaline phosphatase (ALP) were normal. The serum levels of tartrate-resistant acid phosphatase (TRAP), a parameter for bone resorption, were increased (4.6 KAU). The serum level of parathyroid hormone-related protein (PTHrP) was elevated (343 pmol/l). The cytokines with stimulatory effects on bone resorption, such as interleukin (IL)-1alpha, IL-1beta, IL-6, and tumor necrosis factor-alpha, were not detected. Serum Ca levels, PTHrP levels, and TRAP levels decreased with the decrease in ATL cells after chemotherapy, while serum BGP levels and ALP levels increased. On the 29th hospital day, ATL cells began to increase again. Then serum PTHrP levels, Ca levels, and TRAP levels increased, while serum BGP levels and ALP levels decreased. A marked excessive bone resorption with suppressed bone formation (uncoupling) occurred in this patient. The ATL cells produced not only PTHrP but also IL-1alpha and IL-1beta. These results suggest that PTHrP may act as a humoral factor and IL-1 may act as a local factor in bone metabolism of ATL patients.

Bone Resorption↗
PubMed2001

The spinal nerves innervate putative chemosensory cells in the ventral skin of desert toads, Bufo alvarius.

Toads normally obtain water by absorption across their skin from osmotically dilute sources. When hyperosmotic salt solutions are presented as a hydration source to dehydrated desert toads, they place the ventral skin onto the source but soon afterwards escape to avoid dehydration. The escape behavior coincides with neural excitation of the spinal nerves that innervate putative chemosensory cells in the ventral skin. In the present study, fluorescent dye translocated through the spinal nerves to those receptor cells in the epidermis was photoconverted in the presence of 3, 3'-diaminobenzidine tetrahydrochloride for electron-microscopic observation of the cells and associated nerve terminals. Most of the photoconverted cells were located in the deepest layer of the epidermis, with some being in more intermediate layers. No labeled cell was seen in the outermost layer of living cells. In desert toads, flask cells and Merkel cells are occasionally seen in the epidermis. An association of nerve fibers with these epidermal cells has been reported in some species of the anurans. In the present study, however, the cytological features of the photoconverted cells are neither reminiscent of flask cells nor Merkel cells, but are similar to those of surrounding epithelial cells in each layer of the epidermis. We hypothesize a sensory function for these cells, because they have a close association with nerve fibers and participate in the transepithelial transport of salts that must pass through all cell layers of the skin.

Animals↗
PubMed2001

Chromosomal instability detected by fluorescence in situ hybridization in Japanese breast cancer patients.

The relationship between chromosomal instability (CIN) and prognostic factors was investigated in 31 breast cancers and 5 benign breast lesions (three fibroadenomas and two papillomas). Using fluorescence in situ hybridization (FISH) with chromosome-specific DNA probes of chromosomes 1, 2, 6, 7, 10, 11, 17 and 18, CIN for each case was determined. CIN varied from 8.1% to 59.3% among the breast cancer patients tested, and was significantly higher than that observed in the benign breast lesions (p<0.01). Moreover, CIN showed a significant correlation with lymph node metastases (p<0.05) and estrogen receptor negativity (p<0.01). These findings suggest that CIN might be useful in the prediction of the biological aggressiveness of breast cancers.

Asian People↗
PubMed2001

Contribution of myeloperoxidase to coronary artery vasculitis associated with MPO-ANCA production.

The role of myeloperoxidase (MPO) in the pathogenesis of vasculitis associated with MPO-specific anti-neutrophil cytoplasmic autoantibody (MPO-ANCA) was examined in a murine animal model. Coronary artery vasculitis was induced in C57BL/6 mice with and without endogenous MPO by intraperitoneal injection of Candida albicans-derived substances (CADS). The corresponding levels of MPO-ANCA in sera of mice with and without vasculitis were measured and compared in both wild-type and MPO-deficient animals. The MPO-ANCA titers in sera were significantly higher in mice with vasculitis than in vasculitis-negative mice, indicating that MPO-ANCA correlated with vasculitis formation. However, the increase of MPO-ANCA titers observed in sera of wild C57BL/6 mice were strongly suppressed in MPO-deficient C57BL/6 mice, accompanied with prevention of vasculitis formation. These results show that MPO acted as an antigen for MPO-ANCA production by CADS and was followed by the vasculitis formation. Vasculitis did develop in a few MPO-deficient mice, though the incidence of vasculitis was much lower in MPO-deficient mice than in C57BL/6 mice.

Animals↗
PubMed2001

Adenovirus-mediated ex vivo gene transfer of basic fibroblast growth factor promotes collateral development in a rabbit model of hind limb ischemia.

Adenovirus-mediated ex vivo gene transfer of basic fibroblast growth factor (bFGF), a new strategy for the treatment of chronic vascular occlusive disease, was examined in a rabbit model of hind limb ischemia. The left femoral artery was completely excised to induce an ischemic state in the hind limb of male rabbits. Simultaneously, a skin section was resected from the wound, and host fibroblasts were cultured. The cultured fibroblasts were infected with adenovirus vector containing modified human bFGF cDNA with the secretory signal sequence (AxCAMAssbFGF) or LacZ cDNA (AxCALacZ). At 21 days after femoral artery excision, the gene-transduced fibroblasts were administered through the left internal iliac artery. The fibroblasts significantly accumulated in the ischemic hind limb, and the AxCAMAssbFGF-treated cells secreted bFGF for less than 14 days without elevation of systemic bFGF level. At 28 days after cell administration, calf blood pressure ratio, angiographic score, capillary density of muscle tissue and blood flow of the left internal iliac artery were determined, and animals with AxCAMAssbFGF-treated cells showed significantly greater development of collateral vessels, as compared with those with AxCALacZ-treated cells. These findings suggest that adenovirus-mediated ex vivo gene transfer of bFGF was effective for improvement of chronic limb ischemia.

Adenoviridae↗
PubMed2001

PKC1, a protein kinase C homologue of Saccharomyces cerevisiae, participates in microtubule function through the yeast EB1 homologue, BIM1.

BACKGROUND: RSC is a chromatin-remodelling complex of Saccharomyces cerevisiae and essential for growth. Its catalytic subunit is encoded by the NPS1/STH1 gene. At the present time, little is known regarding the cellular function of RSC. RESULTS: To identify genes with functions related to NPS1, we screened high-copy suppressor genes for the temperature- and thiabendazole (TBZ)-sensitive mutant allele of NPS1, nps1-105. Amongst the suppressors we cloned PKC1/STT1 and BIM1 that encoded a homologue of mammalian protein kinase C and a conserved microtubule binding protein homologous to human EB1, respectively. Both the temperature sensitive mutation of PKC1, stt1, and the bim1 null mutation caused synthetic growth defects with nps1-105. A genetic analysis of the functional relationships between these genes revealed that PKC1 suppressed the defect of nps1-105 through the BIM1 function but not by the activation of the MPK1/MAPK pathway. The stt1 mutation alone showed TBZ sensitivity and delayed the G2-phase progression at semi-permissive temperatures. Both of these stt1 phenotypes were suppressed by the over-expression of BIM1. In addition, stt1 as well as nps1-105, mis-segregated a mini-chromosome at frequencies higher than the wild-type at a permissive temperature. The mis-segregation was enhanced in the nps1-105 stt1 double mutant. CONCLUSION: These results suggest that Pkc1p plays a role which is relevant to microtubule functions and that this role is mediated by a hitherto unknown PKC signalling pathway and by Bim1p

Blotting, Northern↗
PubMed2001

Mönckeberg's medial sclerosis and inorganic phosphate in uremia.

Mönckeberg's medial sclerosis (MMS) is one of the characteristic calcified lesions of uremic artery disease and often exhibits osseous metaplasia. Although its pathogenic mechanism is largely unknown, MMS may contain two different pathologic processes: degenerative process leading to apoptosis or necrosis of medial smooth muscle cells and osteogenic process leading to formation of bone-like structures. It has long been known that calcification follows necrosis. Apoptotic/necrotic cells often release matrix vesicles or membranous cellular degradation products resulting from disintegration of the cells that frequently serve as the nidus of calcification. On the other hand, vascular cells may exhibit osteoblastic phenotype in vitro, and some of the markers for osteoblastic differentiation and noncollagenous proteins regulating mineralization have been demonstrated in calcified arterial lesions. As a possible etiologic factor inducing these two responses, hyperphosphatemia among various metabolic disturbances recognized in uremia may play an important role in the development of MMS in uremia.

Animals↗
PubMed2001

Gamma probe and ultrasonographically-guided fine-needle aspiration biopsy of sentinel lymph nodes in breast cancer patients.

AIM: The purpose of the present study was to evaluate the usefulness of gamma probe and ultrasonographically-guided fine-needle aspiration biopsy (FNAB) in the pre-operative detection of sentinel node (SN) metastasis in breast cancer patients. METHODS: Sentinel node biopsy (SNB) was performed in patients with stage I or II breast cancer with clinically negative nodes using dye and radio-isotope. Axillas of 60 patients in whom a hot spot was detected by gamma probe were examined by ultrasonography. Pre-operative diagnosis of SN metastasis by gamma probe and ultrasonographically-guided FNAB was compared with the histological results of SN. RESULTS: The sensitivity, specificity and overall accuracy of ultrasonography in the diagnosis of SN metastasis were 50.0%, 92.1% and 76.7%, respectively. SNs were visualized by ultrasonography in 29 of 60 patients. Of 14 patients with positive results by ultrasonography, four had positive and two had negative cytology. The combination of ultrasonography and ultrasonographically-guided FNAB for visualized nodes had a sensitivity of 78.5%, specificity of 93.3% and overall accuracy of 86.2%. Blind FNAB in the hot spot was not useful in the detection of SN metastasis in patients whose SNs failed to be detected by ultrasonography. CONCLUSIONS: Gamma probe and ultrasonographically-guided FNAB is a potentially useful method for pre-operative detection of SN metastasis. In patients with positive SNs, SNB is not indicated and complete axillary lymph-node dissection can be performed as a primary procedure.

Biopsy, Needle↗
PubMed2001

Application of sentinel node biopsy to gastric cancer surgery.

BACKGROUND: Sentinel node (SN) biopsy has been tried in the management of a variety of cancers with the hope that it would eliminate many unnecessary lymph node dissections, resulting in less morbidity. This important technique, however, has not been tried in gastric cancer surgery. The feasibility of SN biopsy and its accuracy in predicting the lymph node status in patients with gastric cancer were examined in the current study. PATIENTS AND METHODS: SN biopsy was performed in patients with T1 (n = 44) or T2 (n = 30) gastric cancers (ie, immediately after laparotomy, indocyanine green was injected around the primary tumor, and the green-stained nodes [SNs: 2.6 +/- 1.7 nodes per patient] were removed). Then, gastrectomy with extended lymphadenectomy was performed. The unstained nodes (non-SNs: 39 +/- 18 nodes per patient) were obtained from the resected specimens. Both SNs and non-SNs were subjected to histologic examination with hematoxylin-eosin. RESULTS: SNs could be identified in 73 of 74 patients (success rate, 99%). Of these 73 patients, 10 had lymph node metastases in SNs or non-SNs, or both; 6 in both SNs and non-SNs; 3 in SNs alone; and 1 in non-SNs alone. The sensitivity of the SN status in the diagnosis of the lymph node status of the patient was 90% (9/10) and specificity was 100% (63/63). Sensitivity was 100% in the T1 group (n = 44) and 88% in the T2 group (n = 29). CONCLUSIONS: SN biopsy using indocyanine green can be performed with a high success rate, and the SN status can predict the lymph node status with a high degree of accuracy, especially in patients with T1 gastric cancer.

Coloring Agents↗