Showing papers by "Ken-ichi Isobe published in 2010"

Therapeutic angiogenesis by transplantation of induced pluripotent stem cell-derived Flk-1 positive cells

Abstract: Induced pluripotent stem (iPS) cells are the novel stem cell population induced from somatic cells. It is anticipated that iPS will be used in the expanding field of regenerative medicine. Here, we investigated whether implantation of fetal liver kinase-1 positive (Flk-1+) cells derived from iPS cells could improve angiogenesis in a mouse hind limb model of ischemia. Flk-1+ cells were induced from iPS cells after four to five days of culture. Hind limb ischemia was surgically induced and sorted Flk-1+ cells were directly injected into ischemic hind limbs of athymic nude mice. Revascularization of the ischemic hind limb was accelerated in mice that were transplanted with Flk-1+ cells compared with control mice, which were transplanted with vehicle, as evaluated by laser Doppler blood flowmetry. Transplantation of Flk-1+ cells also increased expression of VEGF mRNA in ischemic tissue compared to controls. Direct local implantation of iPS cell-derived Flk-1+ cells would salvage tissues from ischemia. These data indicate that iPS cells could be valuable in the therapeutic induction of angiogenesis.

Toxic effects of D-galactose on thymus and spleen that resemble aging.

Abstract: Continuous low-dose injection of d-galactose induces changes in mice that resemble accelerated aging. As such, these mice have been used as models to study mechanisms of aging. Here, we examined whether repeated (daily, for 60 days) subcutaneous injections (at 50 mg d-galactose/kg) into young adult (i.e., 2-month-old) mice induced changes in key immune system organs that were on par with those associated with aging. The results showed that galactose-treated mice develop histologic changes in their thymic cortical and medullary regions; immunohistochemical analysis revealed unorganized distributions of keratin-5 and keratin-8 proteins in the thymus of these hosts. These histological changes in the thymus of d-galactose-treated mice were also observed in the organs of aged (i.e., 24-month-old control mice); however, in this latter group, these changes were accompanied by a strong infiltration of adipose cells. Galactose-treated mice also evinced alterations within their splenic white and red pulp. Further, ...

Intra-bone marrow bone marrow transplantation rejuvenates the B-cell lineage in aged mice.

Abstract: Age-related reductions in the frequency and absolute number of early B lineage precursors in the bone marrow of aged mice have been reported. Reversal of B-cell lineage senescence has not been achieved. Age-related impairment of the B-cell lineage is caused by the decreasing functionality of hematopoietic and B lineage precursors, and reduced efficacy of bone marrow stromal cells that constitute the bone marrow microenvironment. To induce rejuvenation of aged B cells, we injected whole bone marrow from young donors to irradiated aged recipients through the tibia and analyzed B-cell development and immune responsiveness. In aged mice, we found significant reductions in the frequencies and absolute numbers of pro-B cells (B220(+)CD43(+)CD24(+)BP-1(-) and B220(+)CD43(+)CD24(int)BP-1(+)) and pre-B cells (B220(+)CD43(+)CD24(high)BP-1(+) and B220(+)CD43(-)IgM(-)IgD(-)). Intra-bone marrow bone marrow transplantation (IBM-BMT) of young marrow cells including both hematopoietic stem cells and bone marrow stromal cells reversed the reduction of pro-B cells and pre-B cells. In the periphery, the frequency and absolute number of marginal zone-B cell were not significantly different between young, old and IBM-BMT group. The frequency of follicular-B cells in the IBM-BMT group was significantly increased compared to old group. The frequency of B1a B cells in the peritoneal cavity was significantly decreased in the IBM-BMT group. Antibody production against T-independent antigens was not different among the young, the aged and IBM-BMT groups.

GADD34 suppresses wound healing by upregulating expression of myosin IIA.

Abstract: Wound healing consists of sequential steps of tissue repair, and cell migration is particularly important. In order to analyze the potential function of growth arrest and DNA damage inducible protein 34 (GADD34) in tissue repair, we performed in vitro and in vivo wound healing experiments. In an in vitro scratch assay, GADD34 knockout (KO) mouse embryonic fibroblasts (MEFs) had higher migration rates than did wild type (WT) MEFs. Furthermore, the rate of wound closure was faster in GADD34 KO MEFs than in WT MEFs. Using in vivo punch biopsy assays, GADD34 KO mice had accelerated wound healing compared to WT mice. WT mice expressed higher amounts of myosin IIA in migrating macrophages and myofibroblasts than did GADD34 KO mice. These results indicate that GADD34 negatively regulates cell migration in wound healing via expression of myosin IIA.

Shifts in the balance of brain tryptophan metabolism due to age and systemic administration of lipopolysaccharide

Abstract: The kynurenine (KYN) pathway, which is initi-ated by indoleamine 2, 3-dioxygenase (IDO), is a key tryptophan (TRP) metabolic pathway. It shares TRP with the serotonin (5-HT) pathway. Because activation of the KYN pathway by proinflammatory cytokines induces depressive symptoms, shifts in the balance of TRP metabolism to the KYN pathway are closely related to the etiology of depression. In the present study, the influence of age on the effect of the inflammation response system (IRS) on brain TRP metabolism was investigated. Male ICR mice (PND21) were reared for 4 weeks (younger group) or until they reached 1 year of age (older group), and given an intraperitoneal (i.p.) injection of lipopolysaccharide (LPS). The TRP, KYN, and 5-HT levels were measured in the prefrontal cortex, hippocampus, amygdala, and dorsal raphe nuclei. An increase in TRP and 5-HT levels was observed with age in all brain regions, whereas age was associated with decreases in KYN levels in the dorsal raphe nuclei. In all brain regions, LPS increased TRP levels, while it in-creased KYN levels in the prefrontal cortex and amygdala. Reduced KYN/5-HT ratios in all regions were observed with age, whereas increased KYN/5-HT ratios were observed with LPS in all regions except the dorsal raphe nuclei. Thus, age shifted the balance between the KYN and 5-HT pathways toward the 5-HT pathway, and countered the effects of LPS, which shifted the balance to the KYN pathway. These effects are relevant to the etiology of psychiatric disorders in elderly people.

Method for induction of differentiation of pluripotent stem cell into thymic epithelial cell

Abstract: Disclosed is a means for inducing the differentiation of a pluripotent stem cell into a thymic epithelial progenitor cell or a medullar thymic epithelial cell. The means comprises carrying out a first induction step of causing the differentiation of a pluripotent stem cell into a definitive endodermal cell and a second induction step of causing the differentiation of the definitive endodermal cell into a thymic epithelial progenitor cell or a medullar thymic epithelial cell using FGF8, FGF7, FGF10, BMP4 and lithium chloride in combination, thereby inducing the differentiation of the pluripotent stem cell into the thymic epithelial progenitor cell or the medullar thymic epithelial cell. In the second induction step, a first culture is carried out under conditions in which FGF8 is added but FGF7 and FGF10 are not added to a culture medium and, subsequent to the first culture, a second culture is carried out under conditions in which FGF7 and FGF10 are added to a culture medium.

The wound repair is control by monocyte linage cells

Abstract: One of the most important clinical problems in caring for elderly patients is treatment of pressure ulcers. One component of normal wound healing is the generation of an inflammatory reaction, which is characterized by the sequential infiltration of neutrophils, macrophages and lymphocytes. In aged C57BL/6 mice, wound healing is relatively inefficient. We examined the effects of monocytes lineage cells for wound healing. First, we i.v.transplanted bone marrow cells from GFP mice (C57BL/6 background) to C57BL/6 mice. We found that the rate of wound repair was enhanced by the bone marrow transplantation. By Immunohistochemistry, we can detect GFP positive cells in wound site. By FACS analysis Gr-1+ (neutrophils) and CD11b+ cells (macrophages) are observed in wound tissues. These results indicate that macrophages and/or neutrophils have the capacity to repair wounded tissues. We compare the capacity to repair wound by aging. Although old bone marrow cells also enhanced wound repair by bone marrow transplantation, the rate of wound repair old bone marrow was lower than the transplantation of young bone marrow. These results indicate that wound healing is controlled by monocytes lineage cells including neutrophils and macrophages. These monocytes lineage cells of aged mice have lower capacity to repair wound than those of young mice.

Effect of Aging on Auto-Antibodies to Wounded Tissues

Abstract: Wound healing succeeds tissue destruction. We hypothesized that antibodies might bind to wounded tissues, which would facilitate the engulfment of damaged tissues by macrophages. We detected the autoantibodies in sera of different ages of mice, which bind to wounded tissues. We detected IgG1 binding to wounded tissues by using FITC- labeled anti-IgG1 in C57BL/6 mice. These bands were highest in 4 months old C57BL/6 mice. The sera taken from 2 months and 20 months old mice also bound to wounded tissues, although the bands were weaker than those of 4 months old mice. We also examined the autoreactive IgM binding to wounded tissues. We could detect relatively strong bands even in 2 M old mice. The pattern of these bands was changed by advancing age. Intensity of IgM bands was not decreased by advancing age. The splenectomy reduced the intensity of IgG1 bands especially in 4 months old mice. Although we observed the slight delay of wound repair by splenectomy in 2 M and 20 M old mice, we observed that wound repair was strongly delayed in 4 M old mice. Serum from any age of mice enhanced the macrophage phagocytosis by opsonization. Serum taken from splenectomized mice decreased the opsonizing capacity only at 4 M.

Effect of Aging on Auto-Antibodies to Wounded Tissues~!2010-03-17~!2010-04-30~!2010-06-17~!

Abstract: Wound healing succeeds tissue destruction. We hypothesized that antibodies might bind to wounded tissues, which would facilitate the engulfment of damaged tissues by macrophages. We detected the autoantibodies in sera of different ages of mice, which bind to wounded tissues. We detected IgG1 binding to wounded tissues by using FITC- labeled anti-IgG1 in C57BL/6 mice. These bands were highest in 4 months old C57BL/6 mice. The sera taken from 2 months and 20 months old mice also bound to wounded tissues, although the bands were weaker than those of 4 months old mice. We also examined the autoreactive IgM binding to wounded tissues. We could detect relatively strong bands even in 2 M old mice. The pattern of these bands was changed by advancing age. Intensity of IgM bands was not decreased by advancing age. The splenectomy reduced the intensity of IgG1 bands especially in 4 months old mice. Although we observed the slight delay of wound repair by splenectomy in 2 M and 20 M old mice, we observed that wound repair was strongly delayed in 4 M old mice. Serum from any age of mice enhanced the macrophage phagocytosis by opsonization. Serum taken from splenectomized mice decreased the opsonizing capacity only at 4 M.