95 results on '"Murasawa S"'
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2. Angiotensin II initiates tyrosine kinase Pyk2-dependent signalings leading to activation of Rac1-mediated c-Jun NH2-terminal kinase
- Author
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Murasawa, S., primary
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- 2000
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3. Cardiac-specific overexpression of angiotensin II AT2 receptor causes attenuated response to AT1 receptor-mediated pressor and chronotropic effects.
- Author
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Masaki, H, primary, Kurihara, T, additional, Yamaki, A, additional, Inomata, N, additional, Nozawa, Y, additional, Mori, Y, additional, Murasawa, S, additional, Kizima, K, additional, Maruyama, K, additional, Horiuchi, M, additional, Dzau, V J, additional, Takahashi, H, additional, Iwasaka, T, additional, Inada, M, additional, and Matsubara, H, additional
- Published
- 1998
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4. A film-type photocatalyst incorporating highly active tio2 powder and fluororesin binder: photocatalytic activity and long-term stability
- Author
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Sopyan, I., primary, Watanabe, M., additional, Murasawa, S., additional, Hashimoto, K., additional, and Fujishima, A., additional
- Published
- 1996
- Full Text
- View/download PDF
5. Gene Transcription of Angiotensin II Type 2 Receptor Is Repressed by Growth Factors and Glucocorticoids in PC12 Cells
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Kijima, K., primary, Matsubara, H., additional, Murasawa, S., additional, Maruyama, K., additional, Mori, Y., additional, and Inada, M., additional
- Published
- 1995
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6. Characterization of Glucocorticoid Response Element of Rat Angiotensin II Type 1A Receptor Gene
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Murasawa, S., primary, Matsubara, H., additional, Kanasaki, M., additional, Kijima, K., additional, Maruyama, K., additional, Nio, Y., additional, Okubo, N., additional, Tsukaguchi, H., additional, Mori, Y., additional, and Inada, M., additional
- Published
- 1995
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- View/download PDF
7. Regulation of gene transcription of angiotensin II receptor subtypes in myocardial infarction.
- Author
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Nio, Y, primary, Matsubara, H, additional, Murasawa, S, additional, Kanasaki, M, additional, and Inada, M, additional
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- 1995
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- View/download PDF
8. cAMP responsive element-mediated regulation of the gene transcription of the alpha 1B adrenergic receptor by thyrotropin.
- Author
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Kanasaki, M, primary, Matsubara, H, additional, Murasawa, S, additional, Masaki, H, additional, Nio, Y, additional, and Inada, M, additional
- Published
- 1994
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- View/download PDF
9. Differential gene expression and regulation of angiotensin II receptor subtypes in rat cardiac fibroblasts and cardiomyocytes in culture.
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Matsubara, H, primary, Kanasaki, M, additional, Murasawa, S, additional, Tsukaguchi, Y, additional, Nio, Y, additional, and Inada, M, additional
- Published
- 1994
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- View/download PDF
10. Regulatory elements that mediate expression of the gene for the angiotensin II type 1a receptor for the rat.
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Murasawa, S, primary, Matsubara, H, additional, Urakami, M, additional, and Inada, M, additional
- Published
- 1993
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11. Dose-dependent contribution of CD34-positive cell transplantation to concurrent vasculogenesis and cardiomyogenesis for functional regenerative recovery after myocardial infarction.
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Iwasaki H, Kawamoto A, Ishikawa M, Oyamada A, Nakamori S, Nishimura H, Sadamoto K, Horii M, Matsumoto T, Murasawa S, Shibata T, Suehiro S, and Asahara T
- Published
- 2006
12. Role of calcium-sensitive tyrosine kinase Pyk2/CAKbeta/RAFTK in angiotensin II induced Ras/ERK signaling.
- Author
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Murasawa, Satoshi, Mori, Yasukiyo, Nozawa, Yoshihisa, Masaki, Hiroya, Maruyama, Katsuya, Tsutsumi, Yoshiaki, Moriguchi, Yasutaka, Shibasaki, Yasunobu, Tanaka, Yoko, Iwasaka, Toshiji, Inada, Mitsuo, Matsubara, Hiroaki, Murasawa, S, Mori, Y, Nozawa, Y, Masaki, H, Maruyama, K, Tsutsumi, Y, Moriguchi, Y, and Shibasaki, Y
- Published
- 1998
13. Structure of the rat V1a vasopressin receptor gene and characterization of its promoter region and complete cDNA sequence of the 3'-end.
- Author
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Murasawa, S, Matsubara, H, Kijima, K, Maruyama, K, Mori, Y, and Inada, M
- Abstract
The gene encoding the rat V1a arginine vasopressin (AVP) receptor was isolated, and its structural organization and 5'-flanking region were characterized. In addition, the complete cDNA sequence of the major transcript of the rat V1a receptor gene was determined. Southern blots demonstrated a single copy of the V1a receptor gene in the rat genome, spanning a region of 3.8 kilobases (kb) and consisting of two exons and one intron (1.8 kb). The location of the intron was unique among G protein-coupled receptor genes in that the first exon encodes six of the seven transmembrane regions, the seventh region being encoded by the second exon. Primer extension, RNase protection, and rapid amplification of the 5'-end of the cDNA identified three transcriptional initiation sites (-405, -243, and -237), the major transcription initiation sites being mapped to positions -243 and -237 base pairs (bp) upstream of the ATG initiation codon (+1 bp). This portion of the 5'-flanking region has neither a TATA nor a CCAAT box, is GC-rich but has no GC box motif, and has features of promoters seen in housekeeping genes. Chimeras containing 2.2 kb of the 5'-flanking region and deletion analyses using the chloramphenicol acetyltransferase gene indicated that a "minimal" region, exhibiting promoter activity and tissue specificity, is located between nucleotides -296 and -221, when transfected into vascular smooth muscle cells. Gel mobility shift assay and Southwestern blotting suggested that approximately 30- and approximately 28-kDa nuclear proteins specifically bind to this region. Rapid amplification of the 3'-end of the cDNA showed that the major transcript terminates 442 bp downstream of the stop codon, in agreement with the mRNA size (2.1 kb). This study demonstrated a distinctive feature in the structural organization of the AVP-oxytocin receptor family genes, and characterization of the 5'-flanking region reported here will lead to a better understanding of the mechanism of transcriptional regulation of the rat V1a AVP receptor gene.
- Published
- 1995
14. Identification of a negative cis-regulatory element and trans-acting protein that inhibit transcription of the angiotensin II type 1a receptor gene.
- Author
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Murasawa, S, Matsubara, H, Mori, Y, Kijima, K, Maruyama, K, and Inada, M
- Abstract
The rat angiotensin II type 1a receptor (AT1a-R) gene is expressed in a cell-specific manner. We demonstrated that the negative regulatory element (NRE) between -489 and -331 is active in PC12 cells (Murasawa, S., Matsubara, H., Urakami, M., and Inada, M. (1993) J. Biol. Chem. 268, 26996-27003). Gel retardation assays confirmed that PC12 cells have a trans-acting factor bound to the NRE. By means of a DNase I footprint assay we identified the core of the NRE as an (A+T)-rich sequence (TAATCTTTTATTTTA) located at nucleotides -456 to -442. Oligonucleotides corresponding to the NRE core sequence bound to nuclear protein. Site-directed mutagenesis at nucleotides -451 to -448 eliminated the specific protein/DNA binding and restored expression of the AT1a-R in transient transfection assays (2.7-fold increase). The NRE did not negatively affect the thymidine kinase promoter. No homology was found with known NREs, suggesting that this is a novel NRE. Southwestern blotting revealed a 53-kDa, specific binding protein in PC12 cells and the rat brain, but not in the liver, spleen, adrenal gland, and kidney. These findings demonstrate that the NRE of the rat AT1a-R is an (A+T)-rich sequence located at nucleotides -456 to -442 and the 53-kDa protein is a specific binding protein, and suggest that this protein may be a trans-acting factor which determines the neuron-specific down-regulation of the AT1a-R gene.
- Published
- 1995
15. Numerical modelings of superconducting wires for AC loss calculations
- Author
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Amemiya, N., Murasawa, S.-I., Banno, N., and Miyamoto, K.
- Published
- 1998
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16. Finite element analysis of AC loss in non-twisted Bi-2223 tape carrying AC transport current and/or exposed to DC or AC external magnetic field
- Author
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Amemiya, N., Miyamoto, K., Murasawa, S.-I., Mukai, H., and Ohmatsu, K.
- Published
- 1998
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17. An efficient TiO~2 thin-film photocatalyst: photocatalytic properties in gas-phase acetaldehyde degradation
- Author
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Sopyan, I., Watanabe, M., Murasawa, S., Hashimoto, K., and Fujishima, A.
- Published
- 1996
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18. A film-type photocatalyst incorporating highly active tio 2 powder and fluororesin binder: photocatalytic activity and long-term stability
- Author
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Sopyan, I., Watanabe, M., Murasawa, S., Hashimoto, K., and Fujishima, A.
- Published
- 1996
- Full Text
- View/download PDF
19. A case of successful treatment with repeated segmental embolization for downhill esophageal varices caused by giant goiter.
- Author
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Watanuki Y, Kageyama K, Kakehata S, Murasawa S, Nakada Y, Takayasu S, Sawaya M, Fujita Y, and Daimon M
- Abstract
Downhill esophageal varices often develop because of venous hypertension caused by either superior vena cava obstruction or compression. We herein present a case of downhill esophageal varices caused by a giant goiter in a patient with postoperative Graves' disease. A 66-year-old man presented with an enlarged goiter. Gastrointestinal endoscopy revealed upper esophageal varices. This patient was successfully treated with repeated segmental embolization of the thyroid arteries that fed the goiter, followed by embolization of the inflow vein for downhill esophageal varices. Three years later, no re-enlargement of either the goiter or the appearance of downhill varices was observed. Segmental embolization therapy is thus considered to be a safe alternative for the treatment of downhill esophageal varices caused by giant goiter.
- Published
- 2024
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20. Vasopressin Expressed in Hypothalamic CRF Neurons Causes Impaired Water Diuresis in Secondary Adrenal Insufficiency.
- Author
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Yamagata S, Talukder AH, Murasawa S, Niioka K, Kumagai N, Takagi M, Kawamura M, Natsume R, Abe M, Uchida K, Sato T, Kurose A, Kageyama K, Daimon M, Sakimura K, and Itoi K
- Subjects
- Mice, Animals, Corticotropin-Releasing Hormone genetics, Corticotropin-Releasing Hormone metabolism, Adrenocorticotropic Hormone metabolism, Pituitary Hormone-Releasing Hormones metabolism, Aquaporin 2 genetics, Aquaporin 2 metabolism, Arginine Vasopressin metabolism, Hypothalamus metabolism, Vasopressins metabolism, Paraventricular Hypothalamic Nucleus metabolism, Neurons metabolism, Diuresis, Hyponatremia metabolism, Adrenal Insufficiency
- Abstract
Patients with secondary adrenal insufficiency can present with impaired free water excretion and hyponatremia, which is due to the enhanced secretion of vasopressin (AVP) despite increased total body water. AVP is produced in magnocellular neurons in the paraventricular nucleus of the hypothalamus (PVH) and supraoptic nucleus and in parvocellular corticotropin-releasing factor (CRF) neurons in the PVH. This study aimed to elucidate whether magnocellular AVP neurons or parvocellular CRF neurons coexpressing AVP are responsible for the pathogenesis of hyponatremia in secondary adrenal insufficiency. The number of CRF neurons expressing copeptin, an AVP gene product, was significantly higher in adrenalectomized AVP-floxed mice (AVPfl/fl) than in sham-operated controls. Adrenalectomized AVPfl/fl mice supplemented with aldosterone showed impaired water diuresis under ad libitum access to water or after acute water loading. They became hyponatremic after acute water loading, and it was revealed under such conditions that aquaporin-2 (AQP2) protein levels were increased in the kidney. Furthermore, translocation of AQP2 to the apical membrane was markedly enhanced in renal collecting duct epithelial cells. Remarkably, all these abnormalities observed in the mouse model for secondary adrenal insufficiency were ameliorated in CRF-AVP-/- mice that lacked AVP in CRF neurons. Our study demonstrates that CRF neurons in the PVH are responsible for the pathogenesis of impaired water excretion in secondary adrenal insufficiency., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Endocrine Society.)
- Published
- 2023
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21. Biochemical Evaluation by Confirmatory Tests after Unilateral Adrenalectomy for Primary Aldosteronism.
- Author
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Murasawa S, Kageyama K, Usutani M, Asari Y, Kinoshita N, Nakada Y, Watanuki Y, Takayasu S, and Daimon M
- Subjects
- Humans, Adrenalectomy methods, Aldosterone, Captopril, Furosemide, Renin, Saline Solution, Retrospective Studies, Hyperaldosteronism diagnosis, Hyperaldosteronism surgery, Hypertension
- Abstract
Primary aldosteronism (PA) is the most common cause of endocrine hypertension. Unilateral PA can be cured using unilateral adrenalectomy (Adx). PA surgery outcome (PASO) criteria, which include clinical and biochemical outcomes, have been proposed to evaluate PA cure after Adx. However, clinical outcomes are often inconsistent with biochemical outcomes. In addition, although confirmatory tests are included as endpoints of biochemical outcomes in the PASO criteria, their clinical usefulness has not yet been established. We evaluated clinical parameters and confirmatory test results before and after Adx in 16 patients with PA and assessed the usefulness of the confirmatory tests. The following were the clinical outcomes after Adx: 37.5% complete success, 62.5% partial success, and 0% absent success. The ratio of biochemical complete success was as follows: 69% aldosterone/renin ratio and basal plasma aldosterone concentration, 19% as assessed by the captopril challenge test, 47% as assessed by the saline infusion test, 30% as assessed by the furosemide upright test, and 100% urine aldosterone. Of these, biochemical complete success was judged in four cases by aldosterone/renin ratio and basal plasma aldosterone concentration, one case by captopril challenge test, five cases by saline infusion test, and one case by furosemide upright test. Although clinical outcomes and urine aldosterone levels improved after Adx, confirmatory tests failed to improve in some cases. The current criteria are not considered useful for biochemical evaluation after Adx. To determine whether additional treatment with mineralocorticoid receptor antagonists is required, more accurate biochemical criteria should be established after Adx., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2023 Shingo Murasawa et al.)
- Published
- 2023
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22. Correlation of magnetic resonance images with neuropathology of irreversible metronidazole-induced encephalopathy: an autopsy case report.
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Miki Y, Takeuchi Y, Murasawa S, Takayasu S, Tsushima F, Kakeda S, Mizukami H, and Wakabayashi K
- Subjects
- Female, Humans, Metronidazole adverse effects, Dysarthria, Autopsy, Tremor, Magnetic Resonance Imaging, Diffusion Magnetic Resonance Imaging methods, Wernicke Encephalopathy pathology, Brain Diseases chemically induced, Brain Diseases diagnostic imaging, Pancreatic Neoplasms
- Abstract
Background: Neurological symptoms and radiographic abnormalities may remain in a small proportion of patients with metronidazole-induced encephalopathy (MIE). Although experimental animal models of MIE have suggested a Wernicke's encephalopathy-like pathology, little is known about the histopathological features of MIE. Here we report the first autopsy case of irreversible MIE., Case Presentation: A 72-year-old Japanese woman with pancreatic neuroendocrine tumour and metastatic tumours in the liver developed intraabdominal bleeding from a hepatic abscess. She was administered metronidazole for 79 days (1.5 g/day), which caused dysarthria followed by hand tremor and altered mental status. Brain magnetic resonance imaging at the time of onset revealed hyperintensities in the deep white matter of the bilateral parietal lobes and splenium of the corpus callosum on diffusion-weighted imaging (DWI) with reduced apparent diffusion coefficient (ADC) values. Despite the improvement of dysarthria and hand tremor, her cognition remained affected even after the withdrawal of metronidazole. She died of pancreatic neuroendocrine tumour at the age of 74 years. Histopathological examinations of the brain confirmed a combination of severe demyelination and moderate axonal degeneration, which corresponded to the regions showing abnormal signal intensities on DWI with reduced ADC values. There were no pathological findings suggestive of Wernicke's encephalopathy in the brain., Conclusion: We have demonstrated the clinical, radiographic and histopathological aspects of irreversible MIE. Hyperintensities on DWI with reduced ADC values in affected regions may indicate a poor clinical prognosis due to irreversible pathological damage., (© 2022. The Author(s).)
- Published
- 2022
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23. Growth differentiation factor-15 modulates adrenocorticotropic hormone synthesis in murine AtT-20 corticotroph cells.
- Author
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Kageyama K, Iwasaki Y, Watanuki Y, Murasawa S, Niioka K, Tasso M, Kosugi A, and Daimon M
- Subjects
- Animals, Corticotropin-Releasing Hormone metabolism, Dexamethasone pharmacology, Gene Expression, Humans, Mice, Pro-Opiomelanocortin genetics, Pro-Opiomelanocortin metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Adrenocorticotropic Hormone metabolism, Corticotrophs chemistry, Corticotrophs metabolism, Growth Differentiation Factor 15 metabolism
- Abstract
Growth differentiation factor-15 (GDF15) is a stress-responsive cytokine that plays important roles in regulation of inflammatory responses, cell growth, and cell differentiation. However, the nature of these roles remains unclear. Here, we aimed to examine the regulatory effects of dexamethasone on Gdf15 expression in murine AtT-20 corticotroph cells. Human Gdf15 promoter-driven luciferase reporter constructs were transfected into corticotroph cells to analyze their promoter activity. The effects of time and concentration of dexamethasone on Gdf15 and proopiomelanocortin (Pomc) mRNA levels were assessed using quantitative real-time polymerase chain reaction. Dexamethasone induced Gdf15 transcription and mRNA levels as well as GDF15 production in transfected cells, whereas reduced the Pomc mRNA levels. GDF15 modulated adrenocorticotropic hormone (ACTH) synthesis, and the dexamethasone-mediated reduction in Pomc mRNA levels were partially relieved upon Gdf15 knockdown. We concluded that GDF15 modulated ACTH production in pituitary corticotrophs in an autocrine manner by suppressing Pomc expression and subsequently mediating the negative feedback effect of glucocorticoids, thereby contributing to pituitary stress response and homeostasis., (Copyright © 2022 Elsevier Inc. All rights reserved.)
- Published
- 2022
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24. Eosinophil counts can be a predictive marker of immune checkpoint inhibitor-induced secondary adrenal insufficiency: a retrospective cohort study.
- Author
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Takayasu S, Mizushiri S, Watanuki Y, Yamagata S, Usutani M, Nakada Y, Asari Y, Murasawa S, Kageyama K, and Daimon M
- Subjects
- Adrenal Insufficiency blood, Aged, Biomarkers, C-Reactive Protein analysis, Female, Humans, Leukocyte Count, Longitudinal Studies, Male, Middle Aged, Prognosis, Retrospective Studies, Sodium blood, Adrenal Insufficiency chemically induced, Adrenal Insufficiency immunology, Eosinophils immunology, Immune Checkpoint Inhibitors adverse effects, Neoplasms drug therapy
- Abstract
Immune checkpoint inhibitors (ICIs) treatment can result in endocrine immune-related adverse events (irAEs), including pituitary dysfunction. Quick diagnosis of secondary adrenal insufficiency (AI) is challenging because no universal definition of ICI-induced secondary AI has been agreed. The aim of this study was to clarify the clinical features of ICI-induced secondary AI that can be used for screening in standard clinical practice. This retrospective study was performed using the medical records of patients who received ICIs at Hirosaki University Hospital between 1 September 2014 and 31 January 2021. Longitudinal clinical data of patients who developed AI were analyzed and compared with the data of thyroid irAEs. Regression analysis showed a significant correlation between ICI-induced secondary AI and absolute or relative eosinophil counts at pre-onset of AI, as well as differences or rate of increase in eosinophil counts at baseline and at pre-onset. Absolute eosinophil counts > 198.36/µL or relative eosinophil counts > 5.6% at pre-onset, and a difference of 65.25/µL or a rate of eosinophil count increase of 1.97 between the baseline and at pre-onset showed the best sensitivity and specificity. This is the first report to demonstrate that eosinophil counts can be a predictor of ICI-induced secondary AI., (© 2022. The Author(s).)
- Published
- 2022
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25. [Development of a System for Evaluating the In-room-laser Alignment Including the Horizontality and Verticality Integrated the Light/Radiation Field Coincidence Test and the Winston-Lutz Test].
- Author
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Shimizu H, Murasawa S, Okudaira K, Aoyama T, Anai S, Nishitani K, Kitagawa T, and Sasaki K
- Subjects
- Humans, Particle Accelerators, Phantoms, Imaging, Lasers, Software
- Abstract
Purpose: The in-room laser which is used for patient positioning in radiotherapy is generally projected on the radiation isocenter determined by the Winston-Lutz test and so on. In this study, a couch-mounted verification device was developed that could evaluate all in-room lasers' alignment including the horizontality and verticality at one time. The device has the function to perform the light/radiation field coincidence test and the Winston-Lutz test at the same time. The aim of this report was to introduce the verification procedure for two tests, using the newly developed software and device, and to present the tuning flow of the in-room laser. Moreover, the analysis accuracy of the developed software was evaluated in comparison with commercial software., Methods: First, the light/radiation field was evaluated by using tungsten markers on the central surface of the device. Next, after aligning the long-carved lines on the front and sides of the device with the in-room lasers, the Winston-Lutz test was carried out by using the tungsten sphere in the center of the device. The acquired images were collectively analyzed using the developed software equipped with the reporting function. Additionally, the result of this Winston-Lutz test was compared with the result from commercial software., Results: A series of the light/radiation field coincidence test and the Winston-Lutz test were analyzed using the developed device and software. The results could be easily confirmed using the reporting function of the software. Regarding the result of the Winston-Lutz test, most of the analysis differences between the developed software and commercially available software were within the pixel size (0.22 mm)., Discussions: Since the accuracy of the radiation field affects the result of the Winston-Lutz test, the presented procedure of performing the light/radiation coincidence field test in advance facilitates the interpretation of the error of the Winston-Lutz test. Based on the results of the Winston-Lutz test, we were able to demonstrate the tuning flow of all in-room lasers including the horizontality and verticality by using the developed device., Conclusions: We have developed a couch-mounted verification device and software that can evaluate the light/radiation coincidence field test and the alignment including the horizontality and verticality of the in-room laser used for patient positioning in radiotherapy, and reported its usefulness. The analysis accuracy of the developed software was comparable to that of commercially available software. The use of this device and the developed software would contribute to not only the efficiency of adjusting all in-room lasers' alignment including the horizontality and verticality but also reflect accurately the result of the Winston-Lutz test.
- Published
- 2021
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26. Procedures for the diagnosis of macro-follicle stimulating hormone (FSH) in a patient with high serum FSH concentrations.
- Author
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Chihara K, Hattori N, Matsuda T, Murasawa S, Daimon M, and Shimatsu A
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- Autoantibodies chemistry, Autoantibodies immunology, Female, Humans, Hypoparathyroidism diagnosis, Middle Aged, Molecular Weight, Polyethylene Glycols chemistry, Reagent Kits, Diagnostic, Urea chemistry, Follicle Stimulating Hormone blood, Immunoassay methods
- Published
- 2020
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27. Regulation of gonadotropins by urocortin 2 in gonadotropic tumor LβT2 cells.
- Author
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Kageyama K, Murasawa S, Niioka K, Otsuka F, Yagi H, and Daimon M
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- Animals, Cell Line, Tumor, Follicle Stimulating Hormone metabolism, Luteinizing Hormone metabolism, Mice, Pituitary Neoplasms genetics, RNA, Messenger metabolism, Receptors, LHRH metabolism, Corticotropin-Releasing Hormone metabolism, Gene Expression Regulation, Gonadotropins, Pituitary metabolism, Pituitary Neoplasms metabolism, Receptors, Corticotropin-Releasing Hormone metabolism, Urocortins metabolism
- Abstract
A close interaction has been shown between the hypothalamo-pituitary-gonadal axis and the hypothalamic-pituitary-adrenal axis. Urocortin 2 (Ucn2) has a very high affinity for the corticotropin-releasing factor (CRF) type 2 (CRF
2 ) receptor. Pituitary Ucn2 regulates expression and secretion of gonadotropins in response to stress. The CRF2 receptor in the pituitary contributes to the modulation of gonadotropins. To explore the possible function of Ucn2 and the CRF2 receptor in pituitary gonadotropic tumor cells, we examined the direct regulation of gonadotropins by Ucn2 in a representative pituitary gonadotropic tumor, mouse LβT2 cells. LβT2 cells were found to express CRF1 receptor and CRF2 receptor mRNA. Ucn2 decreased CRF1 receptor mRNA levels, while it increased CRF2 receptor mRNA levels. Ucn2 directly decreased the mRNA levels of both luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in LβT2 cells. Ucn2 also decreased gonadotropin-releasing hormone receptor (GnRHR) mRNA levels. A selective CRF2 receptor antagonist suppressed the Ucn2-induced decreases in LH, FSH, and GnRHR mRNA levels. Ucn2 acts on gonadotrophs expressing the CRF2 receptor, and inhibits the production of gonadotropins in the pituitary gonadotropic tumor cells. (177 words)., (Copyright © 2017 Elsevier B.V. All rights reserved.)- Published
- 2017
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28. Acute mesenteric ischemia and hepatic infarction after treatment of ectopic Cushing's syndrome.
- Author
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Takayasu S, Murasawa S, Yamagata S, Kageyama K, Nigawara T, Watanuki Y, Kimura D, Tsushima T, Sakamoto Y, Hakamada K, Terui K, and Daimon M
- Abstract
Summary: Patients with Cushing's syndrome and excess exogenous glucocorticoids have an increased risk for venous thromboembolism, as well as arterial thrombi. The patients are at high risk of thromboembolic events, especially during active disease and even in cases of remission and after surgery in Cushing's syndrome and withdrawal state in glucocorticoid users. We present a case of Cushing's syndrome caused by adrenocorticotropic hormone-secreting lung carcinoid tumor. Our patient developed acute mesenteric ischemia after video-assisted thoracoscopic surgery despite administration of sufficient glucocorticoid and thromboprophylaxis in the perioperative period. In addition, our patient developed hepatic infarction after surgical resection of the intestine. Then, the patient was supported by total parenteral nutrition. Our case report highlights the risk of microthrombi, which occurred in our patient after treatment of ectopic Cushing's syndrome. Guidelines on thromboprophylaxis and/or antiplatelet therapy for Cushing's syndrome are acutely needed., Learning Points: The present case showed acute mesenteric thromboembolism and hepatic infarction after treatment of ectopic Cushing's syndrome.Patients with Cushing's syndrome are at increased risk for thromboembolic events and increased morbidity and mortality.An increase in thromboembolic risk has been observed during active disease, even in cases of remission and postoperatively in Cushing's syndrome.Thromboprophylaxis and antiplatelet therapy should be considered in treatment of glucocorticoid excess or glucocorticoid withdrawal.
- Published
- 2017
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29. Evaluation of the (1-24) adrenocorticotropin stimulation test for the diagnosis of primary aldosteronism.
- Author
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Terui K, Kageyama K, Nigawara T, Moriyama T, Sakihara S, Takayasu S, Tsushima Y, Watanki Y, Yamagata S, Sugiyama A, Murasawa S, Nakada Y, Suda T, and Daimon M
- Subjects
- Aldosterone blood, Humans, Hydrocortisone blood, Hyperaldosteronism blood, Middle Aged, ROC Curve, Reproducibility of Results, Adrenocorticotropic Hormone pharmacology, Hyperaldosteronism diagnosis, Reagent Kits, Diagnostic
- Abstract
Objective: The purpose of this study was to investigate the diagnostic power of the adrenocorticotropin (ACTH) stimulation test in patients with primary aldosteronism (PA) and those with aldosterone-producing adenoma (APA)., Design: This study was based on a retrospective database analysis., Subjects and Methods: We assessed 158 hypertensive patients with a high plasma aldosterone-to-renin ratio (ARR) including 97 with at least one positive confirmatory test result who did not undergo surgery and comprised a "possible PA" group, 19 with negative results in all tests who were the "non-PA" group, and 41 diagnosed with APA following surgery who were the APA group. The "confirmed PA group" included APA patients and patients from the possible PA group showing both high ARR and hypokalemia. One case was diagnosed as a metastasis., Results: Receiver-operating characteristic (ROC) analysis showed that the diagnostic accuracy of ACTH test was not very effective in differentiating between APA patients and possible PA and non-PA patients. The optimal cut-off value of maximal plasma aldosterone concentration for differentiating between patient in the confirmed PA group and other patients showed moderate accuracy., Conclusions: The ACTH test may not be useful as a screening or confirmatory test, but the test may be useful for differentiating between patients with confirmed PA and the rest of the cohort. The positive finding of the ACTH test may at least support a higher likelihood of lateralizing on adrenal venous sampling., (© The Author(s) 2016.)
- Published
- 2016
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30. The Evaluation of Adrenal Function in Two Cases of Hypocortisolism Accompanied by Liver Cirrhosis.
- Author
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Yamashita M, Kageyama K, Murakami H, Sugiyama A, Yanagimachi M, Sato E, Murasawa S, Matsui J, Tamasawa N, and Daimon M
- Subjects
- Adrenal Insufficiency complications, Adrenal Insufficiency physiopathology, Female, Humans, Liver Cirrhosis complications, Liver Cirrhosis physiopathology, Male, Middle Aged, Adrenal Insufficiency blood, Adrenocorticotropic Hormone blood, Hydrocortisone blood, Liver Cirrhosis blood, Pituitary-Adrenal System physiopathology
- Abstract
Adrenal insufficiency may occur in patients with liver cirrhosis. The assessment of hypothalamus-pituitary-adrenal function is important in such patients, but there is no consensus as to how it should be performed. We herein report the results of our evaluation of the adrenal function in two patients with hypocortisolism accompanied by liver cirrhosis. The patients lacked the typical features of hypocortisolism. One was diagnosed with hypocortisolism accompanied by liver cirrhosis while the other had secondary adrenal insufficiency caused by a hypothalamic disorder. Hypocortisolism accompanied by liver cirrhosis should be evaluated by endocrine tests to determine its pathogenesis. A low-dose adrenocorticotropic hormone test may be appropriate for non-critically ill cirrhotic patients.
- Published
- 2016
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31. Inhibition of heat shock protein 90 decreases ACTH production and cell proliferation in AtT-20 cells.
- Author
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Sugiyama A, Kageyama K, Murasawa S, Ishigame N, Niioka K, and Daimon M
- Subjects
- ACTH-Secreting Pituitary Adenoma metabolism, ACTH-Secreting Pituitary Adenoma pathology, Adenoma metabolism, Adenoma pathology, Adrenocorticotropic Hormone metabolism, Animals, Cell Line, Tumor, Heterocyclic Compounds, 2-Ring pharmacology, Mice, Neoplasm Transplantation, Pro-Opiomelanocortin drug effects, Pro-Opiomelanocortin genetics, Pyrazoles pharmacology, RNA, Messenger metabolism, Real-Time Polymerase Chain Reaction, Securin drug effects, Securin genetics, Tumor Burden, ACTH-Secreting Pituitary Adenoma genetics, Adenoma genetics, Adrenocorticotropic Hormone drug effects, Benzoquinones pharmacology, Cell Proliferation drug effects, HSP90 Heat-Shock Proteins antagonists & inhibitors, Lactams, Macrocyclic pharmacology, RNA, Messenger drug effects
- Abstract
Purpose: Cushing's disease is primarily caused by adrenocorticotropic hormone (ACTH)-producing pituitary adenomas. If excision of the tumor from the pituitary, which is the primary treatment for Cushing's disease, is unsuccessful, further medical therapy is needed to treat the resultant hypercortisolism. Some of the drugs used to treat this condition have shown potential therapeutic benefits, but a more effective treatment should be explored for the treatment of Cushing's disease. In the present study, we determined the effect of heat shock protein 90 inhibitors on ACTH production and cell proliferation of AtT-20 corticotroph tumor cells., Methods: AtT-20 pituitary corticotroph tumor cells were cultured. The expression levels of mouse proopiomelanocortin (POMC) and pituitary tumor transforming gene 1 (PTTG1) mRNA were evaluated using quantitative real-time PCR. Cellular DNA content was analyzed with fluorescence-activated cell sorting (FACS) analysis. The protein levels were determined by Western blot analysis., Results: Both 17-allylamino-17-demethoxygeldanamycin and CCT018159 decreased POMC mRNA levels in AtT-20 cells and ACTH levels in the culture medium of these cells, suggesting that both drugs suppress ACTH synthesis and secretion in corticotroph tumor cells. Both drugs also decreased cell proliferation and induced apoptosis. FACS analyses revealed that both agents increased the percentage of AtT-20 cells in the G2/M phase. These drugs decreased cell proliferation, presumably due to the induction of cell death and arrest of the cell cycle in AtT-20 cells. Tumor weight in mice xenografted with AtT-20 cells and treated with CCT018159 was lower than in AtT-20-xenografted control mice. CCT018159 also decreased plasma ACTH levels, and POMC and PTTG1 mRNA levels in the tumor cells., Conclusions: CCT018159 inhibits ACTH production and corticotroph tumor cell proliferation in vitro and in vivo.
- Published
- 2015
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32. Inhibitory effects of trichostatin A on adrenocorticotropic hormone production and proliferation of corticotroph tumor AtT-20 cells.
- Author
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Nakada Y, Kageyama K, Sugiyama A, Desaki R, Takayasu S, Niioka K, Murasawa S, Ishigame N, Asari Y, Iwasaki Y, and Daimon M
- Subjects
- Adrenocorticotropic Hormone analysis, Animals, Apoptosis drug effects, Cell Line, Tumor, Gene Expression drug effects, Gene Knockdown Techniques, Mice, Pituitary ACTH Hypersecretion, Pro-Opiomelanocortin genetics, RNA, Messenger analysis, Securin genetics, ACTH-Secreting Pituitary Adenoma metabolism, ACTH-Secreting Pituitary Adenoma pathology, Adrenocorticotropic Hormone biosynthesis, Cell Proliferation drug effects, Histone Deacetylase Inhibitors, Hydroxamic Acids pharmacology
- Abstract
Cushing's disease is primarily caused by adrenocorticotropic hormone (ACTH)-producing pituitary adenomas. Pituitary tumor-transforming gene 1 (PTTG1) expression, a hallmark of pituitary tumors, stimulates pituitary cell proliferation. Histone deacetylases (HDACs) play an important role in regulating gene transcription and HDAC inhibitors induce cellular differentiation and suppress tumor cell proliferation. HDAC inhibitors also repress PTTG1 mRNA levels. Trichostatin A (TSA) is a potent cell-permeable HDAC inhibitor that blocks cell cycle progression. In the present study, we determined the effect of TSA on ACTH production and cellular proliferation in mouse AtT-20 corticotroph tumor cells. TSA decreased proopiomelanocortin (POMC) mRNA levels in AtT-20 cells and reduced ACTH levels in the culture medium of these cells. The TSA-induced decreases in POMC mRNA levels were not modulated when TSA and dexamethasone were simultaneously administered. Drug treatment also decreased AtT-20 cell proliferation, induced apoptosis, and increased the percentage of cells in G0/G1 phase using flow cytometry. TSA decreased PTTG1 mRNA levels. Furthermore, PTTG1 knockdown inhibited cellular proliferation. Its knockdown also inhibited POMC mRNA and ACTH levels. TSA inhibits ACTH production and corticotroph tumor cell proliferation. TSA may inhibit cellular proliferation, and ACTH synthesis and secretion by decreasing PTTG1 expression.
- Published
- 2015
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33. Aphidicolin inhibits cell proliferation via the p53-GADD45β pathway in AtT-20 cells.
- Author
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Kageyama K, Sugiyama A, Murasawa S, Asari Y, Niioka K, Oki Y, and Daimon M
- Subjects
- Adrenocorticotropic Hormone metabolism, Animals, Apoptosis drug effects, Apoptosis physiology, Cell Line, Tumor, Cell Proliferation physiology, Mice, Phosphorylation drug effects, Pro-Opiomelanocortin metabolism, Proto-Oncogene Proteins c-akt metabolism, Signal Transduction drug effects, Antigens, Differentiation metabolism, Aphidicolin pharmacology, Cell Proliferation drug effects, Tumor Suppressor Protein p53 metabolism
- Abstract
Cushing's disease is primarily caused by pituitary corticotroph adenomas, which autonomically secrete adrenocorticotropic hormone (ACTH). ACTH production may be associated with tumor cell proliferation; however, the effects of cell cycle progression on ACTH production and cell proliferation are little known in corticotroph tumor cells. A DNA polymerase inhibitor, aphidicolin, arrests cells at the entrance to the S phase and blocks the cell cycle; aphidicolin also induces apoptosis in tumor cells. In the present study, we determined ACTH production and cell proliferation of AtT-20 corticotroph tumor cells following treatment with aphidicolin. Aphidicolin decreased proopiomelanocortin mRNA levels in AtT-20 cells and the levels of ACTH in the culture medium of these cells. Aphidicolin also decreased cell proliferation and induced apoptosis in AtT-20 cells. Fluorescence-activated cell sorting analyses revealed that this agent increased the percentage of G0/G1 phase cells, and decreased S phase cells. Aphidicolin decreased the phosphorylation of cyclic adenosine monophosphate response element-binding protein and Akt. Aphidicolin increased the levels of tumor protein 27 (p27) and 53 (p53), while it decreased cyclin E levels. Aphidicolin also increased the mRNA levels of the stress response gene growth arrest and DNA damage-inducible 45β (GADD45β), a putative downstream target of p53. The p53 knockdown increased GADD45β mRNA levels. The GADD45β knockdown inhibited the decreases in cell proliferation. Thus, aphidicolin inhibits cell proliferation via the p53-GADD45β pathway in AtT-20 cells.
- Published
- 2015
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34. The inhibition of N-glycosylation of glycoprotein 130 molecule abolishes STAT3 activation by IL-6 family cytokines in cultured cardiac myocytes.
- Author
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Matsuo R, Morihara H, Mohri T, Murasawa S, Takewaki K, Nakayama H, Maeda M, and Fujio Y
- Subjects
- Animals, Animals, Newborn, Cells, Cultured, Cytokines metabolism, Extracellular Signal-Regulated MAP Kinases metabolism, Glycosylation, Microscopy, Fluorescence, Rats, Receptors, Interleukin-6 chemistry, Signal Transduction, Cytokine Receptor gp130 metabolism, Interleukin-6 chemistry, Myocytes, Cardiac metabolism, STAT3 Transcription Factor metabolism, Tunicamycin chemistry
- Abstract
Interleukin-6 (IL-6) family cytokines play important roles in cardioprotection against pathological stresses. IL-6 cytokines bind to their specific receptors and activate glycoprotein 130 (gp130), a common receptor, followed by further activation of STAT3 and extracellular signal-regulated kinase (ERK)1/2 through janus kinases (JAKs); however the importance of glycosylation of gp130 remains to be elucidated in cardiac myocytes. In this study, we examined the biological significance of gp130 glycosylation using tunicamycin (Tm), an inhibitor of enzyme involved in N-linked glycosylation. In cardiomyocytes, the treatment with Tm completely replaced the glycosylated form of gp130 with its unglycosylated one. Tm treatment inhibited leukemia inhibitory factor (LIF)-mediated activation of STAT3 and ERK1/2. Similarly, IL-11 failed to activate STAT3 and ERK1/2 in the presence of Tm. Interestingly, Tm inhibited the activation of JAKs 1 and 2, without influencing the expression of suppressor of cytokine signalings (SOCSs) and protein-tyrosine phosphatase 1B (PTP1B), which are endogenous inhibitors of JAKs. To exclude the possibility that Tm blocks LIF and IL-11 signals by inhibiting the glycosylation of their specific receptors, we investigated whether the stimulation with IL-6 plus soluble IL-6 receptor (sIL-6R) could transduce their signals in Tm-treated cardiomyocytes and found that this stimulation was unable to activate the downstream signals. Collectively, these findings indicate that glycosylation of gp130 is essential for signal transduction of IL-6 family cytokines in cardiomyocytes.
- Published
- 2014
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35. Inhibitory effects of SOM230 on adrenocorticotropic hormone production and corticotroph tumor cell proliferation in vitro and in vivo.
- Author
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Murasawa S, Kageyama K, Sugiyama A, Ishigame N, Niioka K, Suda T, and Daimon M
- Subjects
- ACTH-Secreting Pituitary Adenoma genetics, ACTH-Secreting Pituitary Adenoma metabolism, ACTH-Secreting Pituitary Adenoma pathology, Adrenocorticotropic Hormone biosynthesis, Animals, Cell Line, Tumor, Cell Proliferation drug effects, Corticotrophs metabolism, Corticotrophs pathology, Humans, Male, Mice, Mice, Nude, Neoplasm Transplantation, Pituitary Gland metabolism, Pituitary Gland pathology, Pituitary Neoplasms genetics, Pituitary Neoplasms metabolism, Pituitary Neoplasms pathology, Pro-Opiomelanocortin genetics, Pro-Opiomelanocortin metabolism, Proto-Oncogene Proteins c-akt genetics, Proto-Oncogene Proteins c-akt metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, RNA, Small Interfering genetics, RNA, Small Interfering metabolism, Receptors, Somatostatin antagonists & inhibitors, Receptors, Somatostatin genetics, Receptors, Somatostatin metabolism, Somatostatin pharmacology, Tumor Burden drug effects, ACTH-Secreting Pituitary Adenoma drug therapy, Adrenocorticotropic Hormone antagonists & inhibitors, Corticotrophs drug effects, Pituitary Gland drug effects, Pituitary Neoplasms drug therapy, Somatostatin analogs & derivatives
- Abstract
Adrenocorticotropic hormone (ACTH) production by pituitary corticotroph adenomas is the main cause of Cushing's disease. A drug that targets pituitary ACTH-secreting adenomas would aid treatment of Cushing's disease. Octreotide, a somatostatin receptor type 2 (SSTR2)-preferring somatostatin analogue, has no effect on ACTH secretion in patients with Cushing's disease. The multiligand SOM230 (pasireotide) displays a much higher affinity for SSTR1 and SSTR5 than octreotide and suppresses ACTH secretion in cultures of human corticotroph tumors to a greater extent than octreotide. In the present in vitro and in vivo study, we determined the effect of SOM230 on ACTH production and cell proliferation of AtT-20 corticotroph tumor cells. SOM230 decreased proopiomelanocortin (POMC) mRNA levels in AtT-20 cells and ACTH levels in the culture medium of these cells, suggesting that SOM230 suppresses ACTH synthesis and secretion in corticotroph tumor cells. SOM230 also decreased cell proliferation and both cyclic adenosine monophosphate response element-binding protein and Akt phosphorylation in AtT-20 cells. SSTR5 knockdown inhibited the SOM230-induced decreases in cell proliferation. Fluorescence-activated cell sorting analyses revealed that SOM230 did not attenuate cell cycle progression. Tumor weight in mice xenografted with AtT-20 cells and treated with SOM230 was significantly lower than in AtT-20-xenografted control mice. SOM230 also significantly decreased plasma ACTH levels, and POMC and pituitary tumor transforming gene mRNA levels in the tumor cells. Thus, SOM230 inhibits ACTH production and corticotroph tumor cell proliferation in vitro and in vivo., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)
- Published
- 2014
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36. Therapeutic superiority for cartilage repair by CD271-positive marrow stromal cell transplantation.
- Author
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Mifune Y, Matsumoto T, Murasawa S, Kawamoto A, Kuroda R, Shoji T, Kuroda T, Fukui T, Kawakami Y, Kurosaka M, and Asahara T
- Subjects
- Adult, Animals, Apoptosis, Cartilage Diseases metabolism, Cell Differentiation, Chondrogenesis, Collagen Type II genetics, Collagen Type II metabolism, Disease Models, Animal, Female, HLA Antigens metabolism, Humans, Immunohistochemistry, Mesenchymal Stem Cells metabolism, Phenotype, Rats, Rats, Nude, SOX9 Transcription Factor genetics, SOX9 Transcription Factor metabolism, Transplantation, Heterologous, Bone Marrow Cells cytology, Cartilage Diseases therapy, Mesenchymal Stem Cell Transplantation, Mesenchymal Stem Cells cytology, Nerve Tissue Proteins metabolism, Receptors, Nerve Growth Factor metabolism
- Abstract
Recent reports indicated that human isolated CD271+ bone marrow mesenchymal stromal cells (BM-MSCs) have a greater expansion and potential for multipotent differentiation including chondrogenesis than classical plastic adherent (PA) BM-MSCs in vitro. Therefore, we set up a hypothesis that CD271+ MSCs may have a greater chondrogenic potential than PA-MSCs in vitro and in vivo. We investigated the superiority of CD271+ MSCs on chondrogenesis using in vitro expansion and pellet culture system and in vivo rat model of cartilage defect when compared to PA-MSCs. In the in vitro study, CD271+ MSCs showed higher expansion potential and produced larger pellets with higher expressions of chondrogenic genes when compared to the control groups. During the culture, CD271 expression decreased, which resulted in decreased chondrogenesis. In the in vivo study, immunohistochemical staining demonstrated differentiated human chondrocytes identified as double-stained cells with human-specific collagen type 2 and human leukocyte antigen-ABC in CD271+ and PA groups. The number of double-stained cells was significantly higher in the CD271+ group than PA group. Real-time RT-PCR analysis of tissue RNA isolated from the chondral defect site for human-specific chondrogenic markers demonstrated a significantly higher expression in CD271+ group than PA group. Macroscopic examination of chondral defect sites at week 8 revealed glossy white and well-integrated repaired tissues in the CD271+ and PA groups, but not in the PBS group. The average histological score in the CD271+ group was significantly greater than in the other groups. Apoptosis analysis at the cell transplanted site with TUNEL staining showed that the CD271+ group had significantly fewer apoptotic chondrocytes compared with the PA group. These results indicate that CD271+ MSCs have a greater chondrogenic potential than PA-MSCs in both in vitro and in vivo conditions.
- Published
- 2013
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37. Small-molecular inhibitors of Ca²⁺-induced mitochondrial permeability transition (MPT) derived from muscle relaxant dantrolene.
- Author
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Murasawa S, Iuchi K, Sato S, Noguchi-Yachide T, Sodeoka M, Yokomatsu T, Dodo K, Hashimoto Y, and Aoyama H
- Subjects
- Calcium metabolism, Crystallography, X-Ray, Cyclosporine chemical synthesis, Cyclosporine chemistry, Cyclosporine pharmacology, Cyclosporins chemical synthesis, Cyclosporins chemistry, Cyclosporins pharmacology, Dantrolene analogs & derivatives, Dantrolene chemistry, Dose-Response Relationship, Drug, HL-60 Cells, Humans, Mitochondria drug effects, Mitochondria metabolism, Mitochondrial Membrane Transport Proteins metabolism, Mitochondrial Permeability Transition Pore, Models, Molecular, Molecular Structure, Molecular Weight, Small Molecule Libraries chemical synthesis, Small Molecule Libraries chemistry, Structure-Activity Relationship, Tumor Cells, Cultured, Calcium chemistry, Dantrolene pharmacology, Mitochondrial Membrane Transport Proteins antagonists & inhibitors, Muscle Relaxation drug effects, Small Molecule Libraries pharmacology
- Abstract
A structure consisting of substituted hydantoin linked to a 5-(halophenyl)furan-2-yl group via an amide bond was identified as a promising scaffold for development of low-molecular-weight therapeutic agents to treat vascular dysfunction, including ischemia/reperfusion injury. Among the compounds synthesized, 5-(3,5-dichlorophenyl)-N-{2,4-dioxo-3-[(pyridin-3-yl)methyl]imidazolidin-1-yl}-2-furamide (17) possessed the most potent inhibitory activity against Ca(2+)-induced mitochondrial swelling. The structural development, synthesis and structure-activity relationship of these compounds are described., (Copyright © 2012 Elsevier Ltd. All rights reserved.)
- Published
- 2012
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38. Action of glucagon-like peptide 1 and glucose levels on corticotropin-releasing factor and vasopressin gene expression in rat hypothalamic 4B cells.
- Author
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Kageyama K, Yamagata S, Akimoto K, Sugiyama A, Murasawa S, and Suda T
- Subjects
- Analysis of Variance, Animals, Cells, Cultured, Corticotropin-Releasing Hormone metabolism, Gene Expression Regulation, Glucagon-Like Peptide-1 Receptor, Glucose metabolism, Hypothalamo-Hypophyseal System metabolism, Pituitary-Adrenal System metabolism, Promoter Regions, Genetic, Rats, Receptors, Glucagon genetics, Receptors, Glucagon metabolism, Vasopressins metabolism, Corticotropin-Releasing Hormone genetics, Gene Expression, Glucagon-Like Peptide 1 physiology, Glucose physiology, Hypothalamus cytology, Vasopressins genetics
- Abstract
Corticotropin-releasing factor (CRF) and arginine vasopressin (AVP) are the two major regulatory peptides in the hypothalamic-pituitary-adrenal (HPA) axis. Glucagon-like peptide-1 (GLP-1), an important regulator of metabolism or energy homeostasis, is implicated in the regulation of the HPA axis in response to stress and may act directly on CRF and AVP neurons. To elucidate the direct regulation of CRF and AVP genes by GLP-1 in the hypothalamus, we examined the effect of GLP-1 in hypothalamic 4B cells, which show the characteristics of hypothalamic paraventricular nucleus neurons. The mRNA of GLP-1 receptor was detected in 4B cells by RT-PCR. GLP-1 significantly stimulated both CRF and AVP mRNA levels. Cells were transfected with CRF or AVP promoter to examine the activity of each promoter. GLP-1 directly stimulated the activities of both CRF and AVP promoters in hypothalamic 4B cells. Basal promoter activities of both CRF and AVP were increased in higher glucose medium. In addition, CRF and AVP promoter activities were increased by GLP-1 in standard or low glucose medium but not in higher glucose medium. An equimolar concentration of metabolically inactive l-glucose failed to mimic the effect of d-glucose, indicating that the event was caused by changes in glucose levels and not by hyperosmolality. Together, these data suggest that GLP-1 would contribute to stress responses through activation of CRF and AVP genes in the hypothalamic cells. Hyperglycemia may be one of the stressors enhancing the syntheses of CRF and AVP in the hypothalamus., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)
- Published
- 2012
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39. Therapeutic administration of IL-11 exhibits the postconditioning effects against ischemia-reperfusion injury via STAT3 in the heart.
- Author
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Obana M, Miyamoto K, Murasawa S, Iwakura T, Hayama A, Yamashita T, Shiragaki M, Kumagai S, Miyawaki A, Takewaki K, Matsumiya G, Maeda M, Yoshiyama M, Nakayama H, and Fujio Y
- Subjects
- Animals, Animals, Newborn, Apoptosis drug effects, Cells, Cultured, Cytoprotection, Disease Models, Animal, Dose-Response Relationship, Drug, Gene Expression Regulation, Hemodynamics drug effects, Humans, Injections, Intravenous, Metallothionein genetics, Metallothionein metabolism, Mice, Mice, Inbred C57BL, Mice, Knockout, Myocardial Infarction genetics, Myocardial Infarction metabolism, Myocardial Infarction pathology, Myocardial Infarction physiopathology, Myocardial Reperfusion Injury genetics, Myocardial Reperfusion Injury metabolism, Myocardial Reperfusion Injury pathology, Myocardial Reperfusion Injury physiopathology, Myocardium pathology, Oxidative Stress drug effects, Phosphorylation, RNA Interference, Rats, Rats, Wistar, Reactive Oxygen Species metabolism, STAT3 Transcription Factor deficiency, STAT3 Transcription Factor genetics, Time Factors, Transfection, Ventricular Function, Left drug effects, Ventricular Pressure drug effects, Cardiotonic Agents administration & dosage, Interleukin-11 administration & dosage, Myocardial Infarction prevention & control, Myocardial Reperfusion Injury prevention & control, Myocardium metabolism, STAT3 Transcription Factor metabolism, Signal Transduction drug effects
- Abstract
Activation of cardiac STAT3 by IL-6 cytokine family contributes to cardioprotection. Previously, we demonstrated that IL-11, an IL-6 cytokine family, has the therapeutic potential to prevent adverse cardiac remodeling after myocardial infarction; however, it remains to be elucidated whether IL-11 exhibits postconditioning effects. To address the possibility that IL-11 treatment improves clinical outcome of recanalization therapy against acute myocardial infarction, we examined its postconditioning effects on ischemia/reperfusion (I/R) injury. C57BL/6 mice were exposed to ischemia (30 min) and reperfusion (24 h), and IL-11 was intravenously administered at the start of reperfusion. I/R injury mediated the activation of STAT3, which was enhanced by IL-11 administration. IL-11 treatment reduced I/R injury, analyzed by triphenyl tetrazolium chloride staining [PBS, 46.7 ± 14.4%; IL-11 (20 μg/kg), 28.6 ± 7.5% in the ratio of infarct to risk area]. Moreover, echocardiographic and hemodynamic analyses clarified that IL-11 treatment preserved cardiac function after I/R. Terminal deoxynucleotide transferase-mediated dUTP nick-end labeling staining revealed that IL-11 reduced the frequency of apoptotic cardiomyocytes after I/R. Interestingly, IL-11 reduced superoxide production assessed by in situ dihydroethidium fluorescence analysis, accompanied by the increased expression of metallothionein 1 and 2, reactive oxygen species (ROS) scavengers. Importantly, with the use of cardiac-specific STAT3 conditional knockout (STAT3 CKO) mice, it was revealed that cardiac-specific ablation of STAT3 abrogated IL-11-mediated attenuation of I/R injury. Finally, IL-11 failed to suppress the ROS production after I/R in STAT3 CKO mice. IL-11 administration exhibits the postconditioning effects through cardiac STAT3 activation, suggesting that IL-11 has the clinical therapeutic potential to prevent I/R injury in heart.
- Published
- 2012
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40. Dexamethasone stimulates the expression of ghrelin and its receptor in rat hypothalamic 4B cells.
- Author
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Kageyama K, Akimoto K, Yamagata S, Sugiyama A, Murasawa S, Watanuki Y, Tamasawa N, and Suda T
- Subjects
- Animals, Cells, Cultured, Dose-Response Relationship, Drug, Ghrelin metabolism, Hypothalamus metabolism, RNA, Messenger genetics, RNA, Messenger metabolism, Rats, Real-Time Polymerase Chain Reaction, Receptors, Ghrelin metabolism, Structure-Activity Relationship, Dexamethasone pharmacology, Ghrelin genetics, Hypothalamus cytology, Hypothalamus drug effects, Receptors, Ghrelin genetics
- Abstract
Growth hormone (GH)-releasing peptides (GHRPs) are synthetic peptides that strongly induce GH release. GHRPs act via a specific receptor, the GHRP receptor (GHSR), of which ghrelin is a natural ligand. GHRPs also induce adrenocorticotropic hormone (ACTH) release in healthy subjects. GHRPs or ghrelin stimulate ACTH release via corticotropin-releasing factor (CRF) and arginin vasopressin in the hypothalamus. Stress-activated CRF neurons are suppressed by glucocorticoids in the hypothalamic paraventricular nucleus (PVN), while CRF gene is up-regulated by glucocorticoids in the PVN cells without the influence of input neurons. However, little is known about the regulation of ghrelin and GHSR type 1a (GHSR1a) genes by glucocorticoids in PVN cells. To elucidate the regulation of ghrelin and GHSR gene expression by glucocorticoids in PVN cells, here we used a homologous PVN neuronal cell line, hypothalamic 4B, because these cells show characteristics of the parvocellular neurons of the PVN. These cells also express ghrelin and GHSR1a mRNA. Dexamethasone increased ghrelin mRNA levels. A potent glucocorticoid receptor antagonist, RU-486, significantly blocked dexamethasone-induced increases in ghrelin mRNA levels. Dexamethasone also significantly stimulated GHSR1a mRNA and protein levels. Finally, ghrelin increased CRF mRNA levels, as did dexamethasone. Incubation with both dexamethasone and ghrelin had an additive effect on CRF and ghrelin mRNA levels. The ghrelin-GHSR1a system is activated by glucocorticoids in the hypothalamic cells., (Copyright © 2011 Elsevier B.V. All rights reserved.)
- Published
- 2012
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41. Cardiac differentiation of P19CL6 cells by oxytocin.
- Author
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Fathi F, Murasawa S, Hasegawa S, Asahara T, Kermani AJ, and Mowla SJ
- Subjects
- Animals, Cell Aggregation drug effects, Cell Differentiation drug effects, Dimethyl Sulfoxide pharmacology, Embryonal Carcinoma Stem Cells physiology, Free Radical Scavengers pharmacology, Green Fluorescent Proteins genetics, Mice, Myocardial Contraction, Myocytes, Cardiac physiology, Embryonal Carcinoma Stem Cells cytology, Embryonal Carcinoma Stem Cells drug effects, Myocytes, Cardiac cytology, Oxytocin pharmacology
- Abstract
Background: It has been reported that P19 embryonal carcinoma (EC) cells differentiate into beating cardiomyocytes under the action of oxytocin (OT). It has been suggested that dimethylsulfoxide (DMSO) acts via the oxytocin/oxytocin receptor pathway because an oxytocin receptor antagonist not only blocks oxytocin-induced cardiomyocyte differentiation, but also blocks DMSO-induced differentiation. In this study, the differentiation ability of OT was tested using P19CL6 cells., Methods: P19CL6 cells were cultured as a confluent monolayer and aggregated cells. OT was then added to culture media as an inducing agent. The cells treated with 1% DMSO were used as a positive control group. Differentiated cells were evaluated morphologically and immunocytochemically, as well as by RT-PCR. In addition, a stable line of green fluorescent protein (GFP)-expressing P19CL6 cells were differentiated into beating cardiomyocytes by OT., Results: Aggregated P19CL6 cells could be differentiated into cardiomyocytes, whereas monolayer cells could not differentiate and express specific cardiac muscle marker genes. In the control group, both aggregates and monolayer cells could be differentiated into cardiomyocytes by DMSO. In addition, GFP-expressing P19CL6 cells differentiated efficiently into beating cardiomyocytes when treated with OT. The results of all evaluations confirmed that the differentiated cells were cardiomyocytes., Conclusions: We concluded that embryoid body formation (cell aggregation) is necessary for the differentiation of P19CL6 cells into cardiomyocytes when using OT as an inducer agent. Furthermore, because of the high rate of differentiation efficiency, GFP-expressing cardiomyocytes derived from P19CL6 cells have the potential to be used for regenerative therapies in experimental models.
- Published
- 2009
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42. Changing modified regions in the genome in hematopoietic stem cell differentiation.
- Author
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Hamada T, Murasawa S, Yokoyama A, Hayashi S, Kobayashi Y, and Asahara T
- Subjects
- AC133 Antigen, AMP-Activated Protein Kinases genetics, Antigens, CD analysis, Cloning, Molecular, Glycoproteins analysis, Humans, Microfilament Proteins genetics, Microtubule-Associated Proteins genetics, Peptides analysis, DNA Methylation, Gene Expression, Genome, Human, Hematopoiesis genetics, Hematopoietic Stem Cells physiology
- Abstract
In the process of hematopoietic stem cell (CD133+ cell) differentiation, a drastic change in gene expression occurs which must be regulated by epigenetic mechanisms. One strategy for CD133+ cell differentiation analysis is to identify genomic DNA regions that have been modified in the process of differentiation. However, it is difficult to obtain large amounts of genomic DNA from uniform CD133+ cells. Based on this situation, we screened genomic DNA regions where modifications change during the process of differentiation in human CD133+ cells using differential methylation site scanning (DMSS), which is a method of identifying differentially methylated regions of the genome from a small number of cells. As a result, we cloned three DNA fragments which corresponded to centrosomal protein 68kDA (Cep68), TRIO and F-actin binding protein (TRIOBP), and AMP-activated protein kinase beta (AMPKb).
- Published
- 2009
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43. Cardiogenic potential of endothelial progenitor cells.
- Author
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Murasawa S and Asahara T
- Subjects
- Coronary Vessels pathology, Coronary Vessels physiology, Humans, Regeneration, Endothelial Cells cytology, Hematopoietic Stem Cell Transplantation, Hematopoietic Stem Cells cytology, Myocardial Ischemia therapy
- Abstract
Transplantation of endothelial progenitor cells (EPCs) are one of the promising strategies to recover the capillary flow in ischaemic diseases such as ischaemic heart disease and peripheral artery disease (PAD) in the leg. However, our previous and another group's works suggested the scarcity of the number of EPCs in peripheral blood might cause insufficient effect for the ischaemic diseases. There are several strategies to overcome this issue, such as (1) in vivo EPC expansion; (2) ex vivo EPC expansion; (3) local (not systemic) EPC injection; and (4) modification of EPC by gene transfer. Recent publications from our own and other groups have reported the possibility of cardiogenic potential of EPCs. We would like to focus on the strategies of EPC transplantation and cardiomyogenesis of EPCs in this review.
- Published
- 2008
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44. Functional and gene expression analysis of hTERT overexpressed endothelial cells.
- Author
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Takano H, Murasawa S, and Asahara T
- Abstract
Telomerase dysfunction contributes to cellular senescence. Recent advances indicate the importance of senescence in maintaining vascular cell function in vitro. Human telomerase reverse transcriptase (hTERT) overexpression is thought to lead to resistance to apoptosis and oxidative stress. However, the mechanism in endothelial lineage cells is unclear. We tried to generate an immortal endothelial cell line from human umbilical vein endothelial cells using a no-virus system and examine the functional mechanisms of hTERT overexpressed endothelial cell senescence in vitro. High levels of hTERT genes and endothelial cell-specific markers were expressed during long-term culture. Also, angiogenic responses were observed in hTERT over-expressed endothelial cell. These cells showed a delay in senescence and appeared more resistant to stressed conditions. PI3K/Akt-related gene levels were enhanced in hTERT overexpressed endothelial cells. An up-regulated PI3K/Akt pathway caused by hTERT overexpression might contribute to anti-apoptosis and survival effects in endothelial lineage cells.
- Published
- 2008
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45. Simple screening method for differentially methylated regions of the genome using a small number of cells.
- Author
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Hamada T, Murasawa S, and Asahara T
- Subjects
- Cell Line, HeLa Cells, Humans, Kidney cytology, Chromosome Mapping methods, DNA genetics, DNA Fingerprinting methods, DNA Methylation, Kidney physiology, Polymerase Chain Reaction methods, Sequence Analysis, DNA methods
- Abstract
Genomic DNA methylation is a major epigenetic mechanism controlling the expression of genetic information. Therefore, identifying regions of the genome that are differentially methylated in different cells is a useful strategy for the study of biological phenomena. To date, several useful screening methods have been established for identifying differentially methylated genomic regions. However, it is impossible to use these methods in fields of study in which it is difficult to obtain a large number of uniform cells, because considerable amounts of genomic DNA are required. Given this situation, we developed a method for preparing large genomic DNA from a small number of cells, and a simple and highly sensitive method for screening for differentially methylated sites. Combined, these two methods comprise a simple screening method, which we named "Differential Methylation Site Scanning" (DMSS), for identifying differentially methylated regions of the genome from a small number of cells. Just 10 cells are sufficient for the method described here.
- Published
- 2007
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46. Gene modified cell transplantation for vascular regeneration.
- Author
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Murasawa S and Asahara T
- Subjects
- Animals, Capillaries physiology, Cells, Cultured, Disease Models, Animal, Genetic Therapy methods, Humans, Ischemia physiopathology, Ischemia therapy, Mice, Vascular Diseases, Vascular Endothelial Growth Factor A biosynthesis, Vascular Endothelial Growth Factor Receptor-1 biosynthesis, Blood Vessels physiology, Endothelium, Vascular transplantation, Gene Transfer Techniques, Neovascularization, Physiologic genetics, Regeneration genetics, Stem Cell Transplantation
- Abstract
Cell Transplantation is one of the powerful tools to ameliorate the capillary flow in ischemic condition. EPC (Endothelial Progenitor Cell) was identified in adult peripheral blood and thought to be a suitable candidate for cell transplantation. Also, gene therapy is already promising choice for enhancing angiogenic property. The combination of cell transplantation and gene therapy should be more effective way to regenerate vasculature in ischemic region. Recently, several research reports have come out regarding gene modified cell transplantation. We will mainly focus on the background of EPC, and then gene modified EPC findings in this review.
- Published
- 2007
47. [Vascular endothelium regeneration therapy].
- Author
-
Murasawa S and Asahara T
- Subjects
- Adult, Animals, Cell Differentiation, Combined Modality Therapy, Genetic Therapy methods, Humans, Antigens, CD34, Arteriosclerosis therapy, Cell Transplantation, Diabetic Angiopathies therapy, Endothelial Cells cytology, Myocardial Ischemia therapy, Neovascularization, Physiologic physiology, Regenerative Medicine methods
- Abstract
CD34 positive cells were first defined as endothelial progenitor cells(EPCs) from circulating mononuclear cells in peripheral blood. EPCs have shown to be mobilized from bone marrow by the various factors, incorporate into sites of physiological and pathological neovascularization and differentiate into mature endothelial cells (ECs). Post-natal vasculogenesis has been considered to be involved in neovascularization of adult tissues. Recently, freshly isolated CD34 positive cells transplantation has started as clinical trial for ischemic diseases. In the clinical situation, we should consider the cell number and cell quality derived from the patients who have atherosclerosis background, especially diabetes. Ex vivo expansion or gene modification of EPCs could be the strategies for the next generation cell therapy to overcome these issues.
- Published
- 2006
48. Therapeutic potential of vasculogenesis and osteogenesis promoted by peripheral blood CD34-positive cells for functional bone healing.
- Author
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Matsumoto T, Kawamoto A, Kuroda R, Ishikawa M, Mifune Y, Iwasaki H, Miwa M, Horii M, Hayashi S, Oyamada A, Nishimura H, Murasawa S, Doita M, Kurosaka M, and Asahara T
- Subjects
- Angiogenesis Inhibitors pharmacology, Animals, Cell Movement, Cells, Cultured, Endothelial Cells, Female, Femoral Fractures pathology, Femur pathology, Gene Expression, Humans, Leukocytes, Mononuclear chemistry, Mice, Osteocalcin genetics, RNA, Messenger analysis, Rats, Rats, Nude, Antigens, CD34 analysis, Femoral Fractures therapy, Femur blood supply, Leukocytes, Mononuclear transplantation, Neovascularization, Physiologic drug effects, Osteogenesis genetics
- Abstract
Failures in fracture healing are mainly caused by a lack of vascularization. Adult human circulating CD34+ cells, an endothelial/hematopoietic progenitor-enriched cell population, have been reported to differentiate into osteoblasts in vitro; however, the therapeutic potential of CD34+ cells for fracture healing is still unclear. Therefore, we performed a series of experiments to test our hypothesis that functional fracture healing is supported by vasculogenesis and osteogenesis via regenerative plasticity of CD34+ cells. Peripheral blood CD34+ cells, isolated from total mononuclear cells of adult human volunteers, showed gene expression of osteocalcin in 4 of 20 freshly isolated cells by single cell reverse transcriptase-polymerase chain reaction analysis. Phosphate-buffered saline, mononuclear cells, or CD34+ cells were intravenously transplanted after producing nonhealing femoral fractures in nude rats. Reverse transcriptase-polymerase chain reaction and immunohistochemical staining at the peri-fracture site demonstrated molecular and histological expression of human-specific markers for endothelial cells and osteoblasts at week 2. Functional bone healing assessed by biomechanical as well as radiological and histological examinations was significantly enhanced by CD34+ cell transplantation compared with the other groups. Our data suggest circulating human CD34+ cells have therapeutic potential to promote an environment conducive to neovascularization and osteogenesis in damaged skeletal tissue, allowing the complete healing of fractures.
- Published
- 2006
- Full Text
- View/download PDF
49. Niche-dependent translineage commitment of endothelial progenitor cells, not cell fusion in general, into myocardial lineage cells.
- Author
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Murasawa S, Kawamoto A, Horii M, Nakamori S, and Asahara T
- Subjects
- Animals, Antibody Specificity, Biomarkers, Cell Lineage physiology, Cells, Cultured, Coculture Techniques, Endothelial Cells immunology, Gene Expression Profiling, Humans, Immunohistochemistry, Myoblasts, Cardiac immunology, Myocytes, Smooth Muscle cytology, Myocytes, Smooth Muscle immunology, Rats, Rats, Nude, Reverse Transcriptase Polymerase Chain Reaction, Stem Cells immunology, Cell Fusion, Endothelial Cells cytology, Endothelial Cells transplantation, Myoblasts, Cardiac cytology, Stem Cell Transplantation, Stem Cells cytology
- Abstract
Objective: Previous studies from our laboratory have shown therapeutic potential of ex vivo expanded endothelial progenitor cells (EPCs) for myocardial ischemia. Our purpose was to investigate the mechanisms regulating EPC contribution to myocardial regeneration., Methods and Results: To evaluate niche-dependent expression profiles of EPCs in vitro, we performed coculture using cultured EPCs derived from human peripheral blood and rat cardiac myoblast cell line (H9C2). Reverse-transcription polymerase chain reaction (PCR) disclosed the expression of human-specific cardiac markers as well as human-specific smooth muscle markers. Cytoimmunochemistry presented several cocultured cells stained with human specific cardiac antibody. To prove this translineage differentiation in vivo, human cultured EPCs were injected into nude rat myocardial infarction model. Reverse-transcription PCR as well as immunohistochemistry of rat myocardial samples demonstrated the expression of human specific cardiac, vascular smooth muscle, and endothelial markers. We observed the distribution of colors (Qtracker; Quantum Dot Corp) in coculture to detect the fused cells, and the frequency of cell fusion was <1%., Conclusions: EPCs can contribute to not only vasculogenesis but also myogenesis in the ischemic myocardium in vivo. Transdifferentiation, not cell fusion, is dominant for EPCs commitment to myocardial lineage cells. Ex vivo expanded EPCs transplantation might have enhanced therapeutic potential for myocardial regeneration.
- Published
- 2005
- Full Text
- View/download PDF
50. Endothelial progenitor cells for vasculogenesis.
- Author
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Murasawa S and Asahara T
- Subjects
- Animals, Genetic Therapy, Humans, Ischemia surgery, Ischemia therapy, Stem Cell Transplantation, Endothelial Cells physiology, Neovascularization, Pathologic etiology, Neovascularization, Physiologic physiology, Stem Cells physiology
- Abstract
Postnatal vasculogenesis is considered to be involved in neovascularization of adult tissues, because bone marrow-derived endothelial progenitor cells (EPCs) were isolated from circulating mononuclear cells in peripheral blood and were shown to incorporate into sites of physiological and pathological neovascularization and to differentiate into mature endothelial cells. EPCs might have an attractive potential therapeutic application for cardiovascular ischemic diseases as a novel cell-based strategy mainly via a vasculogenesis mechanism.
- Published
- 2005
- Full Text
- View/download PDF
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