Katsunori Sasaki - Academia.edu (original) (raw)

Papers by Katsunori Sasaki

World Journal of Stem Cells, 2015

To facilitate close contacts between transplanted cardiomyocytes and host skeletal muscle using c... more To facilitate close contacts between transplanted cardiomyocytes and host skeletal muscle using cell fusion mediated by hemagglutinating virus of Japan envelope (HVJ-E) and tissue maceration. Cardiomyocytes (1.5 × 10(6)) from fetal rats were first cultured. After proliferation, some cells were used for fusion with adult muscle fibers using HVJ-E. Other cells were used to create cardiomyocyte sheets (area: about 3.5 cm(2) including 2.1 × 10(6) cells), which were then treated with Nile blue, separated, and transplanted between the latissimus dorsi and intercostal muscles of adult rats with four combinations of HVJ-E and/or NaOH maceration: G1: HVJ-E(+), NaOH(+), Cardiomyocytes(+); G2: HVJ-E(-), NaOH(+), Cardiomyocytes(+); G3: HVJ-E(+), NaOH(-), Cardiomyocytes(+); G4: HVJ-E(-), NaOH(-), Cardiomyocytes(-). At 1 and 2 wk after transplantation, the four groups were compared by detection of beating domains, motion images using moving target analysis software, action potentials, gene expression of MLC-2v and Mesp1 by reverse transcription-polymerase chain reaction, hematoxylin-eosin staining, and immunostaining for cardiac troponin and skeletal myosin. In vitro cardiomyocytes were fused with skeletal muscle fibers using HVJ-E. Cardiomyocyte sheets remained in the primary transplanted sites for 2 wk. Although beating domains were detected in G1, G2, and G3 rats, G1 rats prevailed in the number, size, motion image amplitudes, and action potential compared with G2 and G3 rats. Close contacts were only found in G1 rats. At 1 wk after transplantation, the cardiomyocyte sheets showed adhesion at various points to the myoblast layer in the latissimus dorsi muscle. At 2 wk after transplantation, close contacts were seen over a broad area. Part of the skeletal muscle sarcoplasma seemed to project into the myocardiocyte plasma and some nuclei appeared to share both sarcoplasmas. The present results show that close contacts were acquired and facilitated the beating function, thereby providing a new cellular transplantation method using HVJ-E and NaOH maceration.

Molecular medicine reports

Non-transferrin-bound iron (NTBI) refers to all forms of iron in the plasma that bind to ligands ... more Non-transferrin-bound iron (NTBI) refers to all forms of iron in the plasma that bind to ligands other than transferrin, and is considered to be a marker of iron toxicity. A variety of analytical approaches for measuring NTBI have been reported; however, a clinically relevant level of sensitivity has yet to be achieved. In addition, insufficient values of NTBI in some patients and healthy subjects have led to the assumption that there may be contamination of reagents with background iron. The present study re-evaluated the analytical procedures of the assay with regard to the potential points of iron contamination in each step. NTA and tris carbonatocobaltate (III) solutions were prepared with removal of iron contamination, and then quantification of NTBI was performed. As a result, the sensitivity of the high-performance liquid chromatography (HPLC)-based NTBI method was improved by the successful reduction of background iron contamination. Application of our modified method proved...

Clinica chimica acta; international journal of clinical chemistry, 2014

Iron is an essential metal in the body, but its excessive accumulation causes damage in various o... more Iron is an essential metal in the body, but its excessive accumulation causes damage in various organs through free radical production. Iron homeostasis is therefore tightly regulated. However, when iron balance collapses, such as in prolonged transfusion, transferrin (Tf) is fully saturated and non-Tf-bound iron (NTBI) appears in the serum. Monitoring serum NTBI levels is therefore crucial in the assessment of the clinical status of patients with iron overload, since NTBI is associated with cellular and organ damage. Several methods for NTBI determination have been reported, but these are extremely complicated and very few laboratories can quantify NTBI at present. We established a novel assay system utilizing automated analyzers that are widely used in clinical laboratories for diagnostic testing. In this assay, NTBI is chelated by nitrilotriacetic acid (NTA), after which the iron is reduced and transferred to nitroso-PSAP, a chromogen. The assay shows excellent linearity, reprodu...

Biochimica et biophysica acta, 2015

The fenestrations of liver sinusoidal endothelial cells (LSECs) play important roles in the excha... more The fenestrations of liver sinusoidal endothelial cells (LSECs) play important roles in the exchange of macromolecules, solutes, and fluid between blood and surrounding liver tissues in response to hepatotoxic drugs, toxins, and oxidative stress. As excess iron is a hepatotoxin, LSECs may be affected by excess iron. In this study, we found a novel link between LSEC defenestration and hepatic nerve growth factor (NGF) in iron-overloaded mice. By Western blotting, NGF was highly expressed, whereas VEGF and HGF were not, and hepatic NGF mRNA levels were increased according to digital PCR. Immunohistochemically, NGF staining was localized in hepatocytes, while TrkA, an NGF receptor, was localized in LSECs. Scanning electron microscopy revealed LSEC defenestration in mice overloaded with iron as well as mice treated with recombinant NGF. Treatment with conditioned medium from iron-overloaded primary hepatocytes reduced primary LSEC fenestrations, while treatment with an anti-NGF neutrali...

The FASEB Journal, 2000

To target disseminated tumors in vivo, transgenes [␤-galactosidase gene, green fluorescence prote... more To target disseminated tumors in vivo, transgenes [␤-galactosidase gene, green fluorescence protein (GFP) gene, herpes simplex virus thymidine kinase (HSV-TK)] were conjugated to transferrin (Tf) by a biotin-streptavidin bridging, which is stoichiometrically controllable, and Tf receptor (Tf-R) affinity chromatography, which selects Tf conjugates with intact receptor bindings sites from reacting with the linker. Tf-␤-galactosidase plasmid conjugate thus constructed was specifically transfected to human erythroleukemia cells (K562) via Tf-R without the aid of any lysosomotropic agents. The transfection efficiency of the conjugate was superior to those of lipofection (1% staining) and retroviral vector (5%) and slightly lower than that of adenovirus (70%). The high level of expression with our conjugate was confirmed using other tumor cells (M7609, TMK-1) whereas in normal diploid cells (HEL), which express low levels of Tf-R, expression was negligible. When GFP gene conjugates were systemically administered through the tail vein to nude mice subcutaneously inoculated with tumor, expression of GFP mRNA was found almost exclusively in tumors and to a much lesser extent in muscles, whereas GFP revealed by fluorescence microscopy was detected only in the former. To exploit a therapeutic applicability of this method, suicide gene therapy using Tf-HSV-TK gene conjugate for massively metastasized k562 tumors in severe combined immunedeficient mice was conducted, and a marked prolongation of survival and significant reduction of tumor burden were confirmed. Thus, this method could also be used for gene therapy to disseminated tu-In vivo gene delivery to tumor cells by transferrin-streptavidin-DNA conjugate. FASEB

PLoS ONE, 2010

Background: The hedgehog (Hh) pathway has been implicated in the pathogenesis of cancer including... more Background: The hedgehog (Hh) pathway has been implicated in the pathogenesis of cancer including pancreatic ductal adenocarcinoma (PDAC). Recent studies have suggested that the oncogenic function of Hh in PDAC involves signaling in the stromal cells rather than cell autonomous effects on the tumor cells. However, the origin and nature of the stromal cell type(s) that are responsive to Hh signaling remained unknown. Since Hh signaling plays a crucial role during embryonic and postnatal vasculogenesis, we speculated that Hh ligand may act on tumor vasculature specifically focusing on bone marrow (BM)-derived cells.

Nephrology, 2002

... Lam terragonolobus ugglutinm (LTA) was used for detect-ing the proximal tubules, and Dolichos... more ... Lam terragonolobus ugglutinm (LTA) was used for detect-ing the proximal tubules, and Dolichos biflorus agglutinin (DBA) was used for collecting ducts." Prior to the LTA staining, the specimens were treated in a microwave at 100°C for 5 min with 10 mmol/L citrate ... Bar = 0.5 pm. ...

The Journal of Organic Chemistry, 1998

Pectenotoxins (PTXs) isolated from the scallop Patinopecten yessoensis were shown to be involved ... more Pectenotoxins (PTXs) isolated from the scallop Patinopecten yessoensis were shown to be involved in an episode of diarrhetic shellfish poisoning (DSP). A total of eight analogues (PTX1-PTX7 and PTX10) have been isolated to date, and the structures of four of these analogues (PTX1, PTX2, PTX3, and PTX6) have already been elucidated. Here, we report the characterization of PTX4 and PTX7 as 7-epi-PTX1 and 7-epi-PTX6, respectively, on the basis of NMR data and an acid-catalyzed chemical interconversion. The structures of two new artifacts, PTX8 and PTX9, produced following this treatment are also reported.

Hepatology, 1993

In chronic inflammation it is reported that serum iron is depleted and hepatic iron is increased ... more In chronic inflammation it is reported that serum iron is depleted and hepatic iron is increased because of reticuloendothelial system iron blockade. However, recent studies indicate that hepatic parenchymal cells increase the uptake of transferrin-bound iron after in vivo stimulation with bacterial lipopolysaccharide, suggesting that endotoxemia itself or lipopolysaccharide-induced production of inflammation-related cytokines may also be responsible for this phenomenon. In this study the actions of inflammation-related cytokines on the synthesis of iron-binding proteins (transferrin and ferritin) and transferrin receptor and the uptake of transferrin-bound iron were investigated in a human hepatoblastoma cell line, HepG2, which is the most commonly used cell line for examining the regulation of hepatic protein synthesis by cytokines. The cells were exposed to interleukin-1 beta, interleukin-6 or tumor necrosis factor-alpha separately for 24 hr. In each cytokine treatment group, the level of transferrin, which is secreted into the conditioned medium, was found to be decreased compared with that of untreated cells. On the other hand, the biosynthesis of ferritin was markedly elevated after the same treatment. This increase in ferritin by cytokine treatment was diminished when deferoxamine was used concomitantly to deplete intracellular chelatable iron. After stimulation with interleukin-1 beta, interleukin-6 or tumor necrosis factor-alpha, 59Fe-labeled transferrin uptake into the cells was increased by 36%, 48%, or 18%, respectively, and this uptake was inhibited by the addition of excess unlabeled transferrin. A binding study with 125I-labeled diferric transferrin revealed that the three cytokines increased the number of transferrin receptors on the cell surface by 1.15-fold to 1.35-fold.(ABSTRACT TRUNCATED AT 250 WORDS)

Clinica Chimica Acta, 2011

Deferasirox (DFX) is an oral iron chelator that is used worldwide for the treatment of iron overl... more Deferasirox (DFX) is an oral iron chelator that is used worldwide for the treatment of iron overload. Although serum ferritin level is usually measured as a marker of the efficacy of DFX, we sometimes experienced unexplainable changes in other serum markers for iron. We hypothesized that photometric assays for serum iron (sFe) and unsaturated iron binding capacity (UIBC) might be affected by DFX. Measurement of sFe and UIBC was performed using 4 different assay systems. The samples were prepared by adding 0-300 μM DFX to pooled human serum or 15 randomized human serum samples. In some experiments, DFX-iron complex (DFX-Fe) was prepared by mixing iron ammonium citrate solution and DFX solution. Measurement of sFe was influenced by DFX-Fe, while iron-free DFX showed no effect on the value of sFe; DFX-Fe was measured as sFe, undistinguishable from transferrin-bound iron. On the other hand, measurement of serum UIBC was influenced by DFX itself; DFX might have been bound to iron in the reagent used for the assay, leading to an increase in UIBC values. DFX affected the sFe and UIBC assay systems. We must be careful in observing these markers during iron chelation therapy with DFX.

Cell and Tissue Research, 2013

The major molecular signals of pancreatic exocrine development are largely unknown. We examine th... more The major molecular signals of pancreatic exocrine development are largely unknown. We examine the role of fibroblast growth factor 7 (FGF7) in the final induction of pancreatic amylase-containing exocrine cells from induced-pancreatic progenitor cells derived from human embryonic stem (hES) cells. Our protocol consisted in three steps: Step I, differentiation of definitive endoderm (DE) by activin A treatment of hES cell colonies; Step II, differentiation of pancreatic progenitor cells by re-plating of the cells of Step I onto 24-well plates at high density and stimulation with all-trans retinoic acid; Step III, differentiation of pancreatic exocrine cells with a combination of FGF7, glucagon-like peptide 1 and nicotinamide. The expression levels of pancreatic endodermal markers such as Foxa2, Sox17 and gut tube endoderm marker HNF1β were up-regulated in both Step I and II. Moreover, in Step III, the induced cells expressed pancreatic markers such as amylase, carboxypeptidase A and chymotrypsinogen B, which were similar to those in normal human pancreas. From day 8 in Step III, cells immunohistochemically positive for amylase and for carboxypeptidase A, a pancreatic exocrine cell product, were induced by FGF7. Pancreatic progenitor Pdx1-positive cells were localized in proximity to the amylase-positive cells. In the absence of FGF7, few amylase-positive cells were identified. Thus, our three-step culture protocol for human ES cells effectively induces the differentiation of amylase- and carboxypeptidase-A-containing pancreatic exocrine cells.

Cell and Tissue Research, 2008

The aim of this study was to induce organized layered tissues with characteristics of the urinary... more The aim of this study was to induce organized layered tissues with characteristics of the urinary tract from embryonic stem (ES) cells alone. We seeded embryoid bodies (EBs) originating from mouse ES cells onto mono-layered collagen membranes and cultured them in four different media. Group 1 was grown in a mixed medium of keratinocyte serum-free medium (KSFM) and Medium 199, Group 2 in a mixed medium of KSFM and conditioned medium collected from 3T3 fibroblasts, Group 3 in an EB formation medium (control group), and Group 4 was KSFM only (control group). After 28 days, cultured tissues were transplanted into nude mice. Cultured tissues from Groups 1 and 2 formed four-layered structures comprising a stratified epithelium, a submucosal loose connective tissue layer, a smooth muscle cell layer identified immunohistochemically by α-smooth muscle actin, and a deep loose connective tissue layer identical to the adventitia. Immunohistochemistry showed that the epithelia were positive for cytokeratins. Tissues also expressed uroplakin as detected by reverse transcriptase/polymerase chain reaction. In contrast, specimens from Groups 3 and 4 demonstrated necrotic features. Uroplakin-positive (i.e., urothelium-like) cells 2 developed only in Group 2 in the transplanted culture tissues in nude mice. In addition to inducing organized, layered tissues from mouse ES cells directly in vitro, these findings demonstrate that tissues cultured in KSFM plus conditioned medium from 3T3 fibroblasts differentiate into luminal walls similar to those of urinary tract in vivo. These findings suggest a new approach to urinary tract regeneration.

Cardiovascular Research, 2003

Embryonic stem cell-derived cardiomyocytes are a useful source for cell transplantation into the ... more Embryonic stem cell-derived cardiomyocytes are a useful source for cell transplantation into the heart, as well as for tissue engineering of the extracardiac vascular system. The present study was designed to investigate the survival and contractile function of embryonic stem cell-derived cardiomyocytes around large blood vessels to assess the feasibility of their ectopic use for future engineering of cardiovascular tissues. The mouse embryonic stem cell-derived cardiomyocytes were transplanted into the retroperitoneum of the adult nude mice, and the myocardial tissues that developed were characterized by electrophysiological and histological techniques. Macroscopic and electrophysiological analyses showed spontaneously contracting transplants in the host retroperitoneum 7 and 30 days after transplantation. Immunohistochemistry detected developing cardiomyocytes in the transplants on Day 7, which formed the myocardial tissues. They were positive for cardiac troponin I, cadherin, connexin 43, and proliferating cell nuclear antigen, but negative for alpha-smooth muscle actin. Vascular formation was discernible in the transplant tissues. By Day 30, more mature myocardial tissues had been established in the transplants. Electron microscopic study emphasized that the transplant tissues comprised cardiomyocytes, in which myofibrils with organized sarcomeres were observed. Desmosomes, fasciae adherens and gap junctions were evident in the cellular junctions. The cardiomyocytes derived from the mouse ES cells were demonstrated to be viable and function in the ectopic site of the host retroperitoneum up to Day 30, following a process of proliferation and differentiation. Vascularization and host perfusion beneficial for the survival of the cardiomyocytes occurred in the transplants.

Bioscience, Biotechnology, and Biochemistry, 2009

... Katsunori SASAKI, Yasuko MATSUKURA, Kumiko SHIJIMA, Mika MIYAKE,y Daisuke FUJIWARA, and Yutak... more ... Katsunori SASAKI, Yasuko MATSUKURA, Kumiko SHIJIMA, Mika MIYAKE,y Daisuke FUJIWARA, and Yutaka KONISHI ... Med., 56, 105112 (2002). 10) Santos-Buelga C and Scalbert A, J. Sci. Food Agric., 80, 1094 1117 (2000). ...

Bioscience, Biotechnology, and Biochemistry, 2008

We have previously reported that highly oligomeric procyanidins (HOPC) purified from Jatoba, a So... more We have previously reported that highly oligomeric procyanidins (HOPC) purified from Jatoba, a South American herb, ameliorated experimental autoimmune encephalomyelitis (EAE) in mice. In this present study, we report that symptoms of arthritis were also significantly reduced by administering the Jatoba extract, when compared with the vehicle-alone-treated control. Interferon-gamma (IFN-gamma) production by the splenocytes from mice injected with procyanidins was also dramatically decreased. The oral administration of purified HOPC was significantly more effective in disease prevention than the ethanol (EtOH) extract of Jatoba. Green tea polyphenol administration, however, surprisingly facilitated disease development. Observation of the joint histopathology on whole paws derived from the HOPC-treated mice showed complete abrogation of collagen induced arthritis (CIA), a characteristic of chronic inflammation in the synovial tissue. These results demonstrate that HOPC administration had an inhibitory effect on both chronic arthritis and EAE and that the oral administration of HOPC exerted its effect after the induction of secondary immunity.

Annals of Anatomy - Anatomischer Anzeiger, 2001

To disclose cell-to-cell interaction associated with the defensive mechanism of the peritoneum, t... more To disclose cell-to-cell interaction associated with the defensive mechanism of the peritoneum, the peritoneum was stimulated with lipopolysaccharide (LPS) and analyzed three-dimensionally, ultrastructurally, and immunohistochemically with immunoSEM (scanning electron microscopy). The activated hepatic peritoneal surface demonstrated numerous microvilli with the adhesion molecules ICAM-1 and VCAM-1. They were restricted to villi and peaked at 1.5 microg/g body weight of LPS. Delicate strands appeared moderately and were interwoven among microvilli with increasing LPS. These strands did not express ICAM-1 or VCAM-1, but fibronectin. Leukocytes began to adhere to the peritoneal surface above die value of LPS (2.5 microg). These results suggest that the peritoneal surface gives a defensive sheet for cell-to-cell interaction through adhesion molecules and fibronectin.

American Journal of Hematology, 1993

A recombinant plasmid carrying human transferrln receptor cDNA in reverse orientation downstream ... more A recombinant plasmid carrying human transferrln receptor cDNA in reverse orientation downstream from the human cytomegalovirus immediate early prornoterlenhancer element was introduced into the HUH-7 human hepatoma cell line by lipofection. Cell surface transferrin bindlng and iron uptake from transferrin each decreased by about 50% in stable transfectants bearing Integrated antisense DNA expression vector. Northern blot analysis indicated that the abundance of target transferrin receptor message was not altered by antisense RNA. These results suggest that the antisense transcript interferes with expression of the endogenous transferrin receptor gene at the level of translation.