Ishikawa, Keiko



Research Centers and Institutes, Health Center (Hiyoshi)


Assistant Professor (Non-tenured)/Research Associate (Non-tenured)/Instructor (Non-tenured)

Academic Background 【 Display / hide

  • 2006

    Keio University, 医学部

    University, Graduated

Academic Degrees 【 Display / hide

  • Identification of quiescent LGR5⁺ stem cells in the human colon, Keio University, 2022


Papers 【 Display / hide

  • Identification of Quiescent LGR5<sup>+</sup> Stem Cells in the Human Colon

    Ishikawa K., Sugimoto S., Oda M., Fujii M., Takahashi S., Ohta Y., Takano A., Ishimaru K., Matano M., Yoshida K., Hanyu H., Toshimitsu K., Sawada K., Shimokawa M., Saito M., Kawasaki K., Ishii R., Taniguchi K., Imamura T., Kanai T., Sato T.

    Gastroenterology (Gastroenterology)  163 ( 5 ) 1391 - 1406.e24 2022.11

    ISSN  00165085

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    Background & Aims: In the mouse intestinal epithelium, Lgr5+ stem cells are vulnerable to injury, owing to their predominantly cycling nature, and their progenies de-differentiate to replenish the stem cell pool. However, how human colonic stem cells behave in homeostasis and during regeneration remains unknown. Methods: Transcriptional heterogeneity among colonic epithelial cells was analyzed by means of single-cell RNA sequencing analysis of human and mouse colonic epithelial cells. To trace the fate of human colonic stem or differentiated cells, we generated LGR5-tdTomato, LGR5-iCasase9-tdTomato, LGR5-split-Cre, and KRT20-ERCreER knock-in human colon organoids via genome engineering. p27+ dormant cells were further visualized with the p27-mVenus reporter. To analyze the dynamics of human colonic stem cells in vivo, we orthotopically xenotransplanted fluorescence-labeled human colon organoids into immune-deficient mice. The cell cycle dynamics in xenograft cells were evaluated using 5-ethynyl-2′-deoxyuridine pulse-chase analysis. The clonogenic capacity of slow-cycling human stem cells or differentiated cells was analyzed in the context of homeostasis, LGR5 ablation, and 5-fluorouracil–induced mucosal injury. Results: Single-cell RNA sequencing analysis illuminated the presence of nondividing LGR5+ stem cells in the human colon. Visualization and lineage tracing of slow-cycling LGR5+p27+ cells and orthotopic xenotransplantation validated their homeostatic lineage-forming capability in vivo, which was augmented by 5-FU–induced mucosal damage. Transforming growth factor–β signaling regulated the quiescent state of LGR5+ cells. Despite the plasticity of differentiated KRT20+ cells, they did not display clonal growth after 5-FU–induced injury, suggesting that occupation of the niche environment by LGR5+p27+ cells prevented neighboring differentiated cells from de-differentiating. Conclusions: Our results highlight the quiescent nature of human LGR5+ colonic stem cells and their contribution to post-injury regeneration.

  • Diet-mediated constitutive induction of novel IL-4<sup>+</sup> ILC2 cells maintains intestinal homeostasis in mice

    Cui W., Nagano Y., Morita S., Tanoue T., Yamane H., Ishikawa K., Sato T., Kubo M., Hori S., Taniguchi T., Hatakeyama M., Atarashi K., Honda K.

    Journal of Experimental Medicine (Journal of Experimental Medicine)  220 ( 8 )  2023.08

    ISSN  00221007

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    Group 2 innate lymphoid cells (ILC2s) expressing IL-5 and IL-13 are localized at various mucosal tissues and play critical roles in the induction of type 2 inflammation, response to helminth infection, and tissue repair. Here, we reveal a unique ILC2 subset in the mouse intestine that constitutively expresses IL-4 together with GATA3, ST2, KLRG1, IL-17RB, and IL-5. In this subset, IL-4 expression is regulated by mechanisms similar to but distinct from those observed in T cells and is partly affected by IL-25 signaling. Although the absence of the microbiota had marginal effects, feeding mice with a vitamin B1-deficient diet compromised the number of intestinal IL-4+ ILC2s. The decrease in the number of IL-4+ ILC2s caused by the vitamin B1 deficiency was accompanied by a reduction in IL-25–producing tuft cells. Our findings reveal that dietary vitamin B1 plays a critical role in maintaining interaction between tuft cells and IL-4+ ILC2s, a previously uncharacterized immune cell population that may contribute to maintaining intestinal homeostasis.

  • An organoid-based organ-repurposing approach to treat short bowel syndrome

    Sugimoto S., Kobayashi E., Fujii M., Ohta Y., Arai K., Matano M., Ishikawa K., Miyamoto K., Toshimitsu K., Takahashi S., Nanki K., Hakamata Y., Kanai T., Sato T.

    Nature (Nature)  592 ( 7852 ) 99 - 104 2021.04

    ISSN  00280836

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    The small intestine is the main organ for nutrient absorption, and its extensive resection leads to malabsorption and wasting conditions referred to as short bowel syndrome (SBS). Organoid technology enables an efficient expansion of intestinal epithelium tissue in vitro1, but reconstruction of the whole small intestine, including the complex lymphovascular system, has remained challenging2. Here we generate a functional small intestinalized colon (SIC) by replacing the native colonic epithelium with ileum-derived organoids. We first find that xenotransplanted human ileum organoids maintain their regional identity and form nascent villus structures in the mouse colon. In vitro culture of an organoid monolayer further reveals an essential role for luminal mechanistic flow in the formation of villi. We then develop a rat SIC model by repositioning the SIC at the ileocaecal junction, where the epithelium is exposed to a constant luminal stream of intestinal juice. This anatomical relocation provides the SIC with organ structures of the small intestine, including intact vasculature and innervation, villous structures, and the lacteal (a fat-absorbing lymphatic structure specific to the small intestine). The SIC has absorptive functions and markedly ameliorates intestinal failure in a rat model of SBS, whereas transplantation of colon organoids instead of ileum organoids invariably leads to mortality. These data provide a proof of principle for the use of intestinal organoids for regenerative purposes, and offer a feasible strategy for SBS treatment.

  • Chromosome Engineering of Human Colon-Derived Organoids to Develop a Model of Traditional Serrated Adenoma

    Kenta Kawasaki, Masayuki Fujii, Shinya Sugimoto, Keiko Ishikawa, Mami Matano, Yuki Ohta, Kohta Toshimitsu, Sirirat Takahashi, Naoki Hosoe, Shigeki Sekine, Takanori Kanai, Toshiro Sato.

    Gastroeneterology 158 ( 3 ) 638 - 651 2019.10

    Research paper (scientific journal)

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    Background & aims: Traditional serrated adenomas (TSAs) are rare colorectal polyps with unique histologic features. Fusions in R-spondin genes have been found in TSAs, but it is not clear whether these are sufficient for TSA development, due to the lack of a chromosome engineering platform for human tissues. We studied the effects of fusions in R-spondin genes and other genetic alterations found in TSA using CRISPR-Cas9-mediated chromosome and genetic modification of human colonic organoids.

    Methods: We introduced chromosome rearrangements that involve R-spondin genes into human colonic organoids, with or without disruption of TP53, using CRISPR-Cas9 (chromosome-engineered organoids). We then knocked a mutation into BRAF encoding the V600E substitution and overexpressed the GREM1 transgene; the organoids were transplanted into colons of NOG mice and growth of xenograft tumors was measured. Colon tissues were collected and analyzed by immunohistochemistry or in situ hybridization. We also established 2 patient-derived TSA organoid lines and characterized their genetic features and phenotypes. We inserted a bicistronic cassette expressing a dimerizer-inducible suicide gene and fluorescent marker downstream of the LGR5 gene in the chromosome-engineered organoids; addition of the dimerizer eradicates LGR5+ cells. Some tumor-bearing mice were given intraperitoneal injections of the dimerizer to remove LGR5-expressing cells.

    Results: Chromosome engineering of organoids required disruption of TP53 or culture in medium containing IGF1 and FGF2. In colons of mice, organoids that expressed BRAFV600E and fusions in R-spondin genes formed flat serrated lesions. Patient-derived TSA organoids grew independent of exogenous R-spondin, and 1 line grew independent of Noggin. Organoids that overexpressed GREM1, in addition to BRAFV600E and fusions in R-spondin genes, formed polypoid tumors in mice that had histologic features similar to TSAs. Xenograft tumors persisted after loss of LGR5-expressing cells.

    Conclusions: We demonstrated efficient chromosomal engineering of human normal colon organoids. We introduced genetic and chromosome alterations into human colon organoids found in human TSAs; tumors grown from these organoids in mice had histopathology features of TSAs. This model might be used to study progression of human colorectal tumors with RSPO fusion gene and GREM1 overexpression.

  • Divergent Routes toward Wnt and R-spondin Niche Independency during Human Gastric Carcinogenesis

    Kosaku Nanki, Kohta Toshimitsu, Ai Takano, Masayuki Fujii, Mariko Shimokawa, Yuki Ohta, Mami Matano, Takashi Seino, Shingo Nishikori, Keiko Ishikawa, Kenta Kawasaki, Kazuhiro Togasaki, Sirirat Takahashi, Yasutaka Sukawa, Hiroki Ishida, Shinya Sugimoto, Hirofumi Kawakubo, Jihoon Kim, Yuko Kitagawa, Shigeki Sekine, Bon-Kyoung Koo, Takanori Kanai, Toshiro Sato.

    Cell 174 ( 4 ) 856 - 869 2018.08

    Research paper (scientific journal)

Papers, etc., Registered in KOARA 【 Display / hide