Browsing by Subject "Stem-cell differentiation"

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    Author Correction: Generation of the organotypic kidney structure by integrating pluripotent stem cell-derived renal stroma
    (Springer Nature, 2023-04-04) Tanigawa, Shunsuke; Tanaka, Etsuko; Miike, Koichiro; Ohmori, Tomoko; Inoue, Daisuke; Cai, Chen-Leng; Taguchi, Atsuhiro; Kobayashi, Akio; Nishinakamura, Ryuichi; Pediatrics, School of Medicine
    Correction to: Nature Communications 10.1038/s41467-022-28226-7, published online 01 February 2022
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    eIF2α signaling regulates autophagy of osteoblasts and the development of osteoclasts in OVX mice
    (Springer Nature, 2019-12-09) Li, Jie; Li, Xinle; Liu, Daquan; Hamamura, Kazunori; Wan, Qiaoqiao; Na, Sungsoo; Yokota, Hiroki; Zhang, Ping; Biomedical Engineering, School of Engineering and Technology
    Bone loss in postmenopausal osteoporosis is induced chiefly by an imbalance of bone-forming osteoblasts and bone-resorbing osteoclasts. Salubrinal is a synthetic compound that inhibits de-phosphorylation of eukaryotic translation initiation factor 2 alpha (eIF2α). Phosphorylation of eIF2α alleviates endoplasmic reticulum (ER) stress, which may activate autophagy. We hypothesized that eIF2α signaling regulates bone homeostasis by promoting autophagy in osteoblasts and inhibiting osteoclast development. To test the hypothesis, we employed salubrinal to elevate the phosphorylation of eIF2α in an ovariectomized (OVX) mouse model and cell cultures. In the OVX model, salubrinal prevented abnormal expansion of rough ER and decreased the number of acidic vesiculars. It regulated ER stress-associated signaling molecules such as Bip, p-eIF2α, ATF4 and CHOP, and promoted autophagy of osteoblasts via regulation of eIF2α, Atg7, LC3, and p62. Salubrinal markedly alleviated OVX-induced symptoms such as reduction of bone mineral density and bone volume fraction. In primary bone-marrow-derived cells, salubrinal increased the differentiation of osteoblasts, and decreased the formation of osteoclasts by inhibiting nuclear factor of activated T-cells cytoplasmic 1 (NFATc1). Live cell imaging and RNA interference demonstrated that suppression of osteoclastogenesis is in part mediated by Rac1 GTPase. Collectively, this study demonstrates that ER stress-autophagy axis plays an important role in OVX mice. Bone-forming osteoblasts are restored by maintaining phosphorylation of eIF2α, and bone-resorbing osteoclasts are regulated by inhibiting NFATc1 and Rac1 GTPase.
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    Generation of the organotypic kidney structure by integrating pluripotent stem cell-derived renal stroma
    (Springer Nature, 2022-02-01) Tanigawa, Shunsuke; Tanaka, Etsuko; Miike, Koichiro; Ohmori, Tomoko; Inoue, Daisuke; Cai, Chen-Leng; Taguchi, Atsuhiro; Kobayashi, Akio; Nishinakamura, Ryuichi; Pediatrics, School of Medicine
    Organs consist of the parenchyma and stroma, the latter of which coordinates the generation of organotypic structures. Despite recent advances in organoid technology, induction of organ-specific stroma and recapitulation of complex organ configurations from pluripotent stem cells (PSCs) have remained challenging. By elucidating the in vivo molecular features of the renal stromal lineage at a single-cell resolution level, we herein establish an in vitro induction protocol for stromal progenitors (SPs) from mouse PSCs. When the induced SPs are assembled with two differentially induced parenchymal progenitors (nephron progenitors and ureteric buds), the completely PSC-derived organoids reproduce the complex kidney structure, with multiple types of stromal cells distributed along differentiating nephrons and branching ureteric buds. Thus, integration of PSC-derived lineage-specific stroma into parenchymal organoids will pave the way toward recapitulation of the organotypic architecture and functions.
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    Passage number affects differentiation of sensory neurons from human induced pluripotent stem cells
    (Springer Nature, 2022-09-23) Cantor, Erica L.; Shen, Fei; Jiang, Guanglong; Tan, Zhiyong; Cunningham, Geneva M.; Wu, Xi; Philips, Santosh; Schneider, Bryan P.; Medicine, School of Medicine
    Induced pluripotent stem cells (iPSCs) are a valuable resource for neurological disease-modeling and drug discovery due to their ability to differentiate into neurons reflecting the genetics of the patient from which they are derived. iPSC-derived cultures, however, are highly variable due to heterogeneity in culture conditions. We investigated the effect of passage number on iPSC differentiation to optimize the generation of sensory neurons (iPSC-dSNs). Three iPSC lines reprogrammed from the peripheral blood of three donors were differentiated into iPSC-dSNs at passage numbers within each of the following ranges: low (5-10), intermediate (20-26), and high (30-38). Morphology and pluripotency of the parent iPSCs were assessed prior to differentiation. iPSC-dSNs were evaluated based on electrophysiological properties and expression of key neuronal markers. All iPSC lines displayed similar morphology and were similarly pluripotent across passage numbers. However, the expression levels of neuronal markers and sodium channel function analyses indicated that iPSC-dSNs differentiated from low passage numbers better recapitulated the sensory neuron phenotype than those differentiated from intermediate or high passage numbers. Our results demonstrate that lower passage numbers may be better suited for differentiation into peripheral sensory neurons.
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    Spontaneously Differentiated GATA6-Positive Human Embryonic Stem Cells Represent an Important Cellular Step in Human Embryonic Development; They Are Not Just an Artifact of In Vitro Culture
    (Mary Ann Liebert, Inc., 2013-10-15) Lee, Jun Ho; Hong, Ki Sung; Mantel, Charlie; Broxmeyer, Hal E.; Lee, Man Ryul; Kim, Kye-Seong; Department of Microbiology & Immunology, School of Medicine
    In this study, we isolated and characterized spontaneously differentiated human embryonic stem cells (SD-hESCs) found in hESC colonies in comparison to the morphologically premature ESCs in the colonies to investigate the potential role of SD-hESCs in embryogenesis. SD-hESCs were distinguished from undifferentiated hESCs by their higher expression of GATA6, a marker for primitive endoderm and transthyretin, a marker visceral endoderm in embryoid bodies (EBs). SD-hESCs expressed OCT4 and NANOG, markers for pluripotent stem cells, at significantly lower levels than undifferentiated hESCs. EBs derived from isolated SD-hESCs were morphologically distinct from cells directly derived from the undifferentiated hESCs; they contained higher number of cysts compared to EBs from undifferentiated hESC-derived EBs (42% vs. 20%). Furthermore, the extracellular signal molecule, BMP2/4, induced a higher GATA4/6 expression and cystic EB formation than control and noggin-treated EBs. Since cystic formation in EBs play a role in primitive endoderm formation during embryogenesis, the SD-hESC may be a relevant cell type equipped to differentiate into primitive endoderm. Our results suggest that SD-ESCs generated during routine hESC culture are not just an artifact of in vitro culture and these cells could serve as a useful model to study the process of embryogenesis.
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    Transgenic ferret models define pulmonary ionocyte diversity and function
    (Springer Nature, 2023) Yuan, Feng; Gasser, Grace N.; Lemire, Evan; Montoro, Daniel T.; Jagadeesh, Karthik; Zhang, Yan; Duan, Yifan; Levlev, Vitaly; Wells, Kristen L.; Rotti, Pavana G.; Shahin, Weam; Winter, Michael; Rosen, Bradley H.; Evans, Idil; Cai, Qian; Yu, Miao; Walsh, Susan A.; Acevedo, Michael R.; Pandya, Darpan N.; Akurathi, Vamsidhar; Dick, David W.; Wadas, Thaddeus J.; Joo, Nam Soo; Wine, Jeffrey J.; Birket, Susan; Fernandez, Courtney M.; Leung, Hui Min; Tearney, Guillermo J.; Verkman, Alan S.; Haggie, Peter M.; Scott, Kathleen; Bartels, Douglas; Meyerholz, David K.; Rowe, Steven M.; Liu, Xiaoming; Yan, Ziying; Haber, Adam L.; Sun, Xingshen; Engelhardt, John F.; Medicine, School of Medicine
    Speciation leads to adaptive changes in organ cellular physiology and creates challenges for studying rare cell-type functions that diverge between humans and mice. Rare cystic fibrosis transmembrane conductance regulator (CFTR)-rich pulmonary ionocytes exist throughout the cartilaginous airways of humans1,2, but limited presence and divergent biology in the proximal trachea of mice has prevented the use of traditional transgenic models to elucidate ionocyte functions in the airway. Here we describe the creation and use of conditional genetic ferret models to dissect pulmonary ionocyte biology and function by enabling ionocyte lineage tracing (FOXI1-CreERT2::ROSA-TG), ionocyte ablation (FOXI1-KO) and ionocyte-specific deletion of CFTR (FOXI1-CreERT2::CFTRL/L). By comparing these models with cystic fibrosis ferrets3,4, we demonstrate that ionocytes control airway surface liquid absorption, secretion, pH and mucus viscosity-leading to reduced airway surface liquid volume and impaired mucociliary clearance in cystic fibrosis, FOXI1-KO and FOXI1-CreERT2::CFTRL/L ferrets. These processes are regulated by CFTR-dependent ionocyte transport of Cl- and HCO3-. Single-cell transcriptomics and in vivo lineage tracing revealed three subtypes of pulmonary ionocytes and a FOXI1-lineage common rare cell progenitor for ionocytes, tuft cells and neuroendocrine cells during airway development. Thus, rare pulmonary ionocytes perform critical CFTR-dependent functions in the proximal airway that are hallmark features of cystic fibrosis airway disease. These studies provide a road map for using conditional genetics in the first non-rodent mammal to address gene function, cell biology and disease processes that have greater evolutionary conservation between humans and ferrets.