In Vivo Stem Cell Imaging Principles and Applications

Seongje Hong,Dong-Sung Lee,Geun-Woo Bae,Juhyeong Jeon,Hak Kyun Kim,Siyeon Rhee,Kyung Oh Jung Korean Society for Stem Cell Research 2023 International journal of stem cells Vol.16 No.4

Stem cells are the foundational cells for every organ and tissue in our body. Cell-based therapeutics using stem cells in regenerative medicine have received attracting attention as a possible treatment for various diseases caused by congenital defects. Stem cells such as induced pluripotent stem cells (iPSCs) as well as embryonic stem cells (ESCs), mesenchymal stem cells (MSCs), and neuroprogenitors stem cells (NSCs) have recently been studied in various ways as a cell-based therapeutic agent. When various stem cells are transplanted into a living body, they can differentiate and perform complex functions. For stem cell transplantation, it is essential to determine the suitability of the stem cell-based treatment by evaluating the origin of stem, the route of administration, in vivo bio-distribution, transplanted cell survival, function, and mobility. Currently, these various stem cells are being imaged in vivo through various molecular imaging methods. Various imaging modalities such as optical imaging, magnetic resonance imaging (MRI), ultrasound (US), positron emission tomography (PET), and single-photon emission computed tomography (SPECT) have been introduced for the application of various stem cell imaging. In this review, we discuss the principles and recent advances of in vivo molecular imaging for application of stem cell research.

Cell Sources, Liver Support Systems and Liver Tissue Engineering: Alternatives to Liver Transplantation

Lee, Soo Young,Kim, Han Joon,Choi, Dongho Korean Society for Stem Cell Research 2015 International journal of stem cells Vol.8 No.1

<P>The liver is the largest organ in the body; it has a complex architecture, wide range of functions and unique regenerative capacity. The growing incidence of liver diseases worldwide requires increased numbers of liver transplant and leads to an ongoing shortage of donor livers. To meet the huge demand, various alternative approaches are being investigated including, hepatic cell transplantation, artificial devices and bioprinting of the organ itself. Adult hepatocytes are the preferred cell sources, but they have limited availability, are difficult to isolate, propagate poor and undergo rapid functional deterioration in vitro. There have been efforts to overcome these drawbacks; by improving culture condition for hepatocytes, providing adequate extracellular matrix, co-culturing with extra-parenchymal cells and identifying other cell sources. Differentiation of human stem cells to hepatocytes has become a major interest in the field of stem cell research and has progressed greatly. At the same time, use of decellularized organ matrices and 3 D printing are emerging cutting-edge technologies for tissue engineering, opening up new paths for liver regenerative medicine. This review provides a compact summary of the issues, and the locations of liver support systems and tissue engineering, with an emphasis on reproducible and useful sources of hepatocytes including various candidates formed by differentiation from stem cells.</P>

Regulation of Stem Cell Fate by ROS-mediated Alteration of Metabolism

Ryu, Jung Min,Lee, Hyun Jik,Jung, Young Hyun,Lee, Ki Hoon,Kim, Dah Ihm,Kim, Jeong Yeon,Ko, So Hee,Choi, Gee Euhn,Chai, Ing Ing,Song, Eun Ju,Oh, Ji Young,Lee, Sei-Jung,Han, Ho Jae Korean Society for Stem Cell Research 2015 International journal of stem cells Vol.8 No.1

<P>Stem cells have attracted much attention due to their distinct features that support infinite self-renewal and differentiation into the cellular derivatives of three lineages. Recent studies have suggested that many stem cells both embryonic and adult stem cells reside in a specialized niche defined by hypoxic condition. In this respect, distinguishing functional differences arising from the oxygen concentration is important in understanding the nature of stem cells and in controlling stem cell fate for therapeutic purposes. ROS act as cellular signaling molecules involved in the propagation of signaling and the translation of environmental cues into cellular responses to maintain cellular homeostasis, which is mediated by the coordination of various cellular processes, and to adapt cellular activity to available bioenergetic sources. Thus, in this review, we describe the physiological role of ROS in stem cell fate and its effect on the metabolic regulation of stem cells.</P>

Host-Microbe Interactions Regulate Intestinal Stem Cells and Tissue Turnover in Drosophila

Ji-Hoon Lee Korean Society for Stem Cell Research 2024 International journal of stem cells Vol.17 No.1

With the activity of intestinal stem cells and continuous turnover, the gut epithelium is one of the most dynamic tissues in animals. Due to its simple yet conserved tissue structure and enteric cell composition as well as advanced genetic and histologic techniques, Drosophila serves as a valuable model system for investigating the regulation of intestinal stem cells. The Drosophila gut epithelium is in constant contact with indigenous microbiota and encounters externally introduced "non-self" substances, including foodborne pathogens. Therefore, in addition to its role in digestion and nutrient absorption, another essential function of the gut epithelium is to control the expansion of microbes while maintaining its structural integrity, necessitating a tissue turnover process involving intestinal stem cell activity. As a result, the microbiome and pathogens serve as important factors in regulating intestinal tissue turnover. In this manuscript, I discuss crucial discoveries revealing the interaction between gut microbes and the host's innate immune system, closely associated with the regulation of intestinal stem cell proliferation and differentiation, ultimately contributing to epithelial homeostasis.

The Molecular Nature of Very Small Embryonic-Like Stem Cells in Adult Tissues

Kim, YongHwan,Jeong, Jaeho,Kang, Hyunsook,Lim, Jisun,Heo, Jinbeom,Ratajczak, Janina,Ratajczak, Mariusz Z.,Shin, Dong-Myung Korean Society for Stem Cell Research 2014 International journal of stem cells Vol.7 No.2

<P>Pluripotent stem cells (PSCs) have been considered as the most important cells in regenerative medicine as they are able to differentiate into all types of cells in the human body. PSCs have been established from several sources of embryo tissue or by reprogramming of terminally differentiated adult tissue by transduction of so-called Yamanaka factors (<I>Oct4</I>, <I>Sox2</I>, <I>Klf4</I>, and <I>cMyc</I>). Interestingly, accumulating evidence has demonstrated the residence of PSCs in adult tissue and with the ability to differentiate into multiple types of tissue-committed stem cells (TCSCs). We also recently demonstrated that a population of pluripotent Oct4<SUP>+</SUP> SSEA-1<SUP>+</SUP>Sca-1<SUP>+</SUP>Lin<SUP>−</SUP>CD45<SUP>−</SUP> very small embryonic-like stem cells (VSELs) resides in the adult murine bone marrow (BM) and in other murine tissue. These very small (∼3–6 <I>μ</I>m) cells express pluripotent markers such as Oct4, Nanog, and SSEA-1. VSELs could be specified into several tissue-residing TCSCs in response to tissue/organ injury, and thus suggesting that these cells have a physiological role in the rejuvenation of a pool of TCSCs under steady-state conditions. In this review article, we discuss the molecular nature of the rare population of VSELs which have a crucial role in regulating the pluripotency, proliferation, differentiation, and aging of these cells.</P>

Measuring Glutathione Regeneration Capacity in Stem Cells

Jihye Kim,Yi-Xi Gong,Eui Man Jeong Korean Society for Stem Cell Research 2023 International journal of stem cells Vol.16 No.3

Glutathione (GSH) is a chief cellular antioxidant, affecting stem cell functions. The cellular GSH level is dynamically altered by the redox buffering system and transcription factors, including NRF2. Additionally, GSH is differentially regulated in each organelle. We previously reported a protocol for monitoring the real-time GSH levels in live stem cells using the reversible GSH sensor FreSHtracer. However, GSH-based stem cell analysis needs be comprehensive and organelle-specific. Hence, in this study, we demonstrate a detailed protocol to measure the GSH regeneration capacity (GRC) in living stem cells by measuring the intensities of the FreSHtracer and the mitochondrial GSH sensor MitoFreSHtracer using a high-content screening confocal microscope. This protocol typically analyses the GRC in approximately 4 h following the seeding of the cells onto plates. This protocol is simple and quantitative. With some minor modifications, it can be employed flexibly to measure the GRC for the whole-cell area or just the mitochondria in all adherent mammalian stem cells.

The Calcineurin-Drp1-Mediated Mitochondrial Fragmentation is Aligned with the Differentiation of c-Kit Cardiac Progenitor Cells

Attaur Rahman,Yuhao Li,Nur Izzah Ismail,To-Kiu Chan,Yuzhen Li,Dachun Xu,Hao Zhou,Sang-Bing Ong Korean Society for Stem Cell Research 2023 International journal of stem cells Vol.16 No.2

Objective: The heart contains a pool of c-kit⁺ progenitor cells which is believed to be able to regenerate. The differentiation of these progenitor cells is reliant on different physiological cues. Unraveling the underlying signals to direct differentiation of progenitor cells will be beneficial in controlling progenitor cell fate. In this regard, the role of the mitochondria in mediating cardiac progenitor cell fate remains unclear. Specifically, the association between changes in mitochondrial morphology with the differentiation status of c-kit⁺ CPCs remains elusive. In this study, we investigated the relationship between mitochondrial morphology and the differentiation status of c-kit⁺ progenitor cells. Methods and Results: c-kit⁺ CPCs were isolated from 2-month-old male wild-type FVB mice. To activate differentiation, CPCs were incubated in α-minimal essential medium containing 10 nM dexamethasone for up to 7 days. To inhibit Drp1-mediated mitochondrial fragmentation, either 10 μM or 50 μM mdivi-1 was administered once at Day 0 and again at Day 2 of differentiation. To inhibit calcineurin, either 1 μM or 5 μM ciclosporin-A (CsA) was administered once at Day 0 and again at Day 2 of differentiation. Dexamethasone-induced differentiation of c-kit⁺ progenitor cells is aligned with fragmentation of the mitochondria via a calcineurin-Drp1 pathway. Pharmacologically inhibiting mitochondrial fragmentation retains the undifferentiated state of the c-kit⁺ progenitor cells. Conclusions: The findings from this study provide an alternative view of the role of mitochondrial fusion-fission in the differentiation of cardiac progenitor cells and the potential of pharmacologically manipulating the mitochondria to direct progenitor cell fate.

In vivo Evaluation of Human Embryonic Stem Cells Isolated by 57-C11 Monoclonal Antibody

Kim, Won-Tae,Lee, Hyun Min,Kim, Min Kyu,Choi, Hong Seo,Ryu, Chun Jeih Korean Society for Stem Cell Research 2016 International journal of stem cells Vol.9 No.2

<P><B>Background</B></P><P>The normal cells derived from human embryonic stem cells (hESCs) are regarded as substitutes for damaged or dysfunctional adult cells. However, tumorigenicity of hESCs remains a major challenge in clinical application of hESC-derived cell transplantation. Previously, we generated monoclonal antibody (MAb) 57-C11 specific to the surface molecule on undifferentiated hESCs. The aim of this study is to prove whether 57-C11-positive hESCs are pluripotent and tumorigenic in immunodeficient mice.</P><P><B>Methods</B></P><P>Undifferentiated hESCs were mixed with retinoic acid (RA)-differentiated hESCs at different ratios prior to 57-C11-mediated separation. To isolate 57-C11-positive hESCs from the mixture, biotinylated 57-C11 and streptavidin-coated magnetic beads were added to the mixture. Unbound 57-C11-negative hESCs were first isolated after applying magnet to the cell mixture, and 57-C11-bound hESCs were then released from the magnetic beads. In order to measure the efficiency of separation, 57-C11-positive or -negative hESCs were counted after isolation. To evaluate the efficiency of teratoma formation <I>in vivo</I>, 57-C11-positive or negative cells were further injected into left and right, respectively, testes of nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice.</P><P><B>Results</B></P><P>Approximately 77~100% of undifferentiated hESCs were isolated after applying 57-C11-coated magnetic beads to the mixed cell populations. Importantly, teratomas were not observed in NOD/SCID mice after the injection of isolated 57-C11-negative hESCs, whereas teratomas were observed with 57-C11-positive hESCs.</P><P><B>Conclusion</B></P><P>57-C11-positive hESCs are pluripotent and tumorigenic. The combination of 57-C11 and magnetic beads will be useful to eliminate remaining undifferentiated hESCs for the safe cell transplantation.</P>

Peripheral Neuron-Organoid Interaction Induces Colonic Epithelial Differentiation via Non-Synaptic Substance P Secretion

Young Hyun Che,In Young Choi,Chan Eui Song,Chulsoo Park,Seung Kwon Lim,Jeong Hee Kim,Su Haeng Sung,Jae Hoon Park,Sun Lee,Yong Jun Kim Korean Society for Stem Cell Research 2023 International journal of stem cells Vol.16 No.3

Background and Objectives: The colonic epithelial layer is a complex structure consisting of multiple cell types that regulate various aspects of colonic physiology, yet the mechanisms underlying epithelial cell differentiation during development remain unclear. Organoids have emerged as a promising model for investigating organogenesis, but achieving organ-like cell configurations within colonic organoids is challenging. Here, we investigated the biological significance of peripheral neurons in the formation of colonic organoids. Methods and Results: Colonic organoids were co-cultured with human embryonic stem cell (hESC)-derived peripheral neurons, resulting in the morphological maturation of columnar epithelial cells, as well as the presence of enterochromaffin cells. Substance P released from immature peripheral neurons played a critical role in the development of colonic epithelial cells. These findings highlight the vital role of inter-organ interactions in organoid development and provide insights into colonic epithelial cell differentiation mechanisms. Conclusions: Our results suggest that the peripheral nervous system may have a significant role in the development of colonic epithelial cells, which could have important implications for future studies of organogenesis and disease modeling.

Strategic Application of Epigenetic Regulators for Efficient Neuronal Reprogramming of Human Fibroblasts

Gary Stanley Fernandes,Rishabh Deo Singh,Debojyoti De,Kyeong Kyu Kim Korean Society for Stem Cell Research 2023 International journal of stem cells Vol.16 No.2

Background and Objectives: Cellular reprogramming in regenerative medicine holds great promise for treating patients with neurological disorders. In this regard, small molecule-mediated cellular conversion has attracted special attention because of its ease of reproducibility, applicability, and fewer safety concerns. However, currently available protocols for the direct conversion of somatic cells to neurons are limited in clinical application due of their complex nature, lengthy process, and low conversion efficiency. Methods and Results: Here, we report a new protocol involving chemical-based direct conversion of human fibroblasts (HF) to matured neuron-like cells with a short duration and high conversion efficiency using temporal and strategic dual epigenetic regulation. In this protocol, epigenetic modulation by inhibition of histone deacetylase and bromodomain enabled to overcome "recalcitrant" nature of adult fibroblasts and shorten the duration of neuronal reprogramming. We further observed that an extended epigenetic regulation is necessary to maintain the induced neuronal program to generate a homogenous population of neuron-like cells. Conclusions: Therefore, our study provides a new protocol to produce neurons-like cells and highlights the need of proper epigenetic resetting to establish and maintain neuronal program in HF.