Use Of Pluripotent Stem Cells Research Articles

Therapeutic potential of urine-derived stem cells in renal regeneration following acute kidney injury: A comparative analysis with mesenchymal stem cells.

Acute kidney injury (AKI) is a common clinical syndrome with high morbidity and mortality rates. The use of pluripotent stem cells holds great promise for the treatment of AKI. Urine-derived stem cells (USCs) are a novel and versatile cell source in cell-based therapy and regenerative medicine that provide advantages of a noninvasive, simple, and low-cost approach and are induced with high multidifferentiation potential. Whether these cells could serve as a potential stem cell source for the treatment of AKI has not been determined. To investigate whether USCs can serve as a potential stem cell source to improve renal function and histological structure after experimental AKI. Stem cell markers with multidifferentiation potential were isolated from human amniotic fluid. AKI severe combined immune deficiency (SCID) mice models were induced by means of an intramuscular injection with glycerol. USCs isolated from human-voided urine were administered via tail veins. The functional changes in the kidney were assessed by the levels of blood urea nitrogen and serum creatinine. The histologic changes were evaluated by hematoxylin and eosin staining and transferase dUTP nick-end labeling staining. Meanwhile, we compared the regenerative potential of USCs with bone marrow-derived mesenchymal stem cells (MSCs). Treatment with USCs significantly alleviated histological destruction and functional decline. The renal function was rapidly restored after intravenous injection of 5 × 105 human USCs into SCID mice with glycerol-induced AKI compared with injection of saline. Results from secretion assays conducted in vitro demonstrated that both stem cell varieties released a wide array of cytokines and growth factors. This suggests that a mixture of various mediators closely interacts with their biochemical functions. Two types of stem cells showed enhanced tubular cell proliferation and decreased tubular cell apoptosis, although USC treatment was not more effective than MSC treatment. We found that USC therapy significantly improved renal function and histological damage, inhibited inflammation and apoptosis processes in the kidney, and promoted tubular epithelial proliferation. Our study demonstrated the potential of USCs for the treatment of AKI, representing a new clinical therapeutic strategy.

Mesendodermal cells fail to contribute to heart formation following blastocyst injection.

Blastocyst complementation offers an opportunity for generating transplantable whole organs from donor sources. Pluripotent stem cells (PSCs) have traditionally served as the primary donor cells due to their ability to differentiate into any type of body cell. However, the use of PSCs raises ethical concerns, particularly regarding their uncontrollable differentiation potential to undesired cell lineages such as brain and germline cells. To address this issue, various strategies have been explored, including the use of genetically modified PSCs with restricted lineage potential or lineage-specified progenitor cells as donors. In this study, we tested whether nascent mesendodermal cells (MECs), which appear during early gastrulation, can be used as donor cells. To do this, we induced Bry-GFP+ MECs from mouse embryonic stem cells (ESCs) and introduced them into the blastocyst. While donor ESCs gave rise to various regions of embryos, including the heart, Bry-GFP+ MECs failed to contribute to the host embryos. This finding suggests that MECs, despite being specified from PSCs within a few days, lack the capacity to assimilate into the developing embryo.

Unlocking the Complexity of Neuromuscular Diseases: Insights from Human Pluripotent Stem Cell-Derived Neuromuscular Junctions.

Over the past 20 years, the use of pluripotent stem cells to mimic the complexities of the human neuromuscular junction has received much attention. Deciphering the key mechanisms underlying the establishment and maturation of this complex synapse has been driven by the dual goals of addressing developmental questions and gaining insight into neuromuscular disorders. This review aims to summarise the evolution and sophistication of in vitro neuromuscular junction models developed from the first differentiation of human embryonic stem cells into motor neurons to recent neuromuscular organoids. We also discuss the potential offered by these models to decipher different neuromuscular diseases characterised by defects in the presynaptic compartment, the neuromuscular junction, and the postsynaptic compartment. Finally, we discuss the emerging field that considers the use of these techniques in drug screening assay and the challenges they will face in the future.

PLURIPOTENCY AND PERSPECTIVES OF CELL TECHNOLOGIES

Biology of pluripotency is a modern field of biological science, and at the same time a tool for modeling human morphogenesis in vitro. Pluripotency is the property of cells to self-renew and differentiate into all types of cells of an adult organism, which appears in early embryogenesis in mammals. Pluripotent stem cells (PSCs) have limitless potential in regenerative and translational medicine, which open up perspectives for solving multiple diseases, including hereditary ones. This review describes the characteristics and uniqueness of PSCs, modeling of early human morphogenesis in vitro in blastocyst-like structures and gastruloids, modeling of organogenesis in organoids. Next, we considered the use of PSCs in regenerative medicine with their risks of capability to oncogenicity and immunogenicity in implication of a cell replacement therapy. However, therapeutic approaches using PSCs are still in their infancy and need to be deeply scrutinized.

Cattle Cloning by Somatic Cell Nuclear Transfer.

Cloning by somatic cell Nuclear Transfer (SCNT) is a powerful technology capable of reprograming terminally differentiated cells to totipotency for generating whole animals or pluripotent stem cells for use in cell therapy, drug screening, and other biotechnological applications. However, the broad usage of SCNT remains limited due to its high cost and low efficiency in obtaining live and healthy offspring. In this chapter, we first briefly discuss the epigenetic constraints responsible for the low efficiency of SCNT and current attempts to overcome them. We then describe our bovine SCNT protocol for delivering live cloned calves and addressing basic questions about nuclear reprogramming. Other research groups can benefit from our basic protocol and build up on it to improve SCNT in the future. Strategies to correct or mitigate epigenetic errors (e.g., correcting imprinting loci, overexpression of demethylases, chromatin-modifying drugs) can integrate the protocol described here.

Pluripotent stem cells: a new strategy for the treatment of diabetes

Diabetes is now one of the most prevalent diseases in the world. Over the next twenty years, the number of affected individuals could reach 693 million. Diabetes is a group of chronic carbohydrate metabolism disorders responsible for increased blood glucose levels. It has several effects on the human body, and over the years can even become disabling, greatly impacting the patient’s quality of life. The most common therapy is based on daily and chronic administration of exogenous insulin, but this is associated with a series of dangerous and potentially fatal complications. Unsurprisingly therefore, over time, studies have explored various alternatives for the treatment of diabetes, not involving the use of insulin, primarily transplantation of the whole pancreas and of the islets of Langerhans. However, neither of these solutions can be applied on a large scale, mainly due to the lack of donors. In recent years, a viable alternative to insulin and to transplantation has emerged through exploration of the use of pluripotent stem cells and their properties. In this review we analyze the state of the art in this field, and the advantages and disadvantages of cell therapies used for reconstruction of the functional unit of the pancreas, the islets of Langerhans.

Stem cell based models in congenital hyperinsulinism - perspective on practicalities and possibilities

Brief Summary: This paper discusses the use of Pluripotent Stem Cell (PSC)-derived pancreatic islets (SC-islets) for studying the basic biology, molecular mechanisms and therapeutic potentials of patients with congenital hyperinsulinism (CHI). The availability of CHI patient islets opens new avenues for research and development of new treatments.

Transformational Applications of Human Cardiac Organoids in Cardiovascular Diseases.

Organoid technology has significantly advanced in recent years and revolutionized the field for generation of organs using in vitro systems (a.k.a “organs in a dish”). The use of pluripotent stem cells or tissue derived cells for generating a 3-dimensional culture system to recapitulate the architecture and function of the organ is central in achieving and improving organoid systems. Unlike most organs in the body, very little progress has been made in cardiac organoid due to its structural complexity and vascularization. In this review, we will discuss the current applications of human cardiac organoids for cardiac disease modeling, drug discovery, drug cardiotoxicity testing, and clinical applications.

Induction of Human Natural Killer Cells Under Defined Conditions by Seamless Transition from Maintenance Culture of Pluripotent Stem Cells.

The use of pluripotent stem cells (PSCs) as a source of natural killer cells (NK cells) can improve reproducibility in the analysis of the pathogenesis of NK cell-associated diseases and in the production of off-the-shelf cellular medicines. We have developed a method for the differentiation of NK cells from human PSCs under serum-free and two-dimensional condition. Our method enables the seamless transition from maintenance of PSCs to differentiation of NK cells, without the use of any techniques other than medium exchange and whole culture passage.

Clever Experimental Designs: Shortcuts for Better iPSC Differentiation.

For practical use of pluripotent stem cells (PSCs) for disease modelling, drug screening, and regenerative medicine, the cell differentiation process needs to be properly refined to generate end products with consistent and high quality. To construct and optimize a robust cell-induction process, a myriad of cell culture conditions should be considered. In contrast to inefficient brute-force screening, statistical design of experiments (DOE) approaches, such as factorial design, orthogonal array design, response surface methodology (RSM), definitive screening design (DSD), and mixture design, enable efficient and strategic screening of conditions in smaller experimental runs through multifactorial screening and/or quantitative modeling. Although DOE has become routinely utilized in the bioengineering and pharmaceutical fields, the imminent need of more detailed cell-lineage specification, complex organoid construction, and a stable supply of qualified cell-derived material requires expedition of DOE utilization in stem cell bioprocessing. This review summarizes DOE-based cell culture optimizations of PSCs, mesenchymal stem cells (MSCs), hematopoietic stem cells (HSCs), and Chinese hamster ovary (CHO) cells, which guide effective research and development of PSC-derived materials for academic and industrial applications.

Process parameter development for the scaled generation of stem cell-derived pancreatic endocrine cells.

Diabetes is a debilitating disease characterized by high blood glucose levels. The global prevalence of this disease has been projected to reach 700 million adults by the year 2045. Type 1 diabetes represents about 10% of the reported cases of diabetes. Although islet transplantation can be a highly effective method to treat type 1 diabetes, its widespread application is limited by the paucity of cadaveric donor islets. The use of pluripotent stem cells as an unlimited cell source to generate insulin‐producing cells for implant is a promising alternative for treating diabetes. However, to be clinically relevant, it is necessary to manufacture these stem cell‐derived cells at sufficient scales. Significant advances have been made in differentiation protocols used to generate stem cell‐derived cells capable of reversing diabetes in animal models and for testing in clinical trials. We discuss the potential of both stem cell‐derived pancreatic progenitors and more matured insulin‐producing cells to treat diabetes. We discuss the need for rigorous bioprocess parameter optimization and identify some critical process parameters and strategies that may influence the critical quality attributes of the cells with the goal of facilitating scalable manufacturing of human pluripotent stem cell‐derived pancreatic endocrine cells.

Pluripotent stem cell therapy for retinal diseases.

Pluripotent stem cells (PSCs), which include human embryonic stem cells (hESCs) and induced pluripotent stem cell (iPSC), have been used to study development of disease processes, and as potential therapies in multiple organ systems. In recent years, there has been increasing interest in the use of PSC-based transplantation to treat disorders of the retina in which retinal cells have been functionally damaged or lost through degeneration. The retina, which consists of neuronal tissue, provides an excellent system to test the therapeutic utility of PSC-based transplantation due to its accessibility and the availability of high-resolution imaging technology to evaluate effects. Preclinical trials in animal models of retinal diseases have shown improvement in visual outcomes following subretinal transplantation of PSC-derived photoreceptors or retinal pigment epithelium (RPE) cells. This review focuses on preclinical studies and clinical trials exploring the use of PSCs for retinal diseases. To date, several phase I/II clinical trials in patients with age-related macular degeneration (AMD) and Stargardt disease (STGD1) have demonstrated the safety and feasibility of PSC-derived RPE transplantation. Additional phase I/II clinical trials using PSC-derived RPE or photoreceptor cells for the treatment of AMD, STGD1, and also retinitis pigmentosa (RP) are currently in the pipeline. As this field continues to evolve, additional technologies may enhance PSC-derived cell transplantation through gene-editing of autologous cells, transplantation of more complex cellular structures such as organoids, and monitoring of transplanted cells through novel imaging technologies.

Transcription Factor-Based Strategies to Generate Neural Cell Types from Human Pluripotent Stem Cells.

In the last years, the use of pluripotent stem cells in studies of human biology has grown exponentially. These cells represent an infinite source for differentiation into several human cell types facilitating the investigation on biological processes, functionality of cells, or diseases mechanisms in relevant human models. In the neurobiology field, pluripotent stem cells have been extensively used to generate the main neuronal and glial cells of the brain. Traditionally, protocols following developmental cues have been applied to pluripotent stem cells to drive differentiation toward different cell lineages; however, these protocols give rise to populations with mixed identities. Interestingly, new protocols applying overexpression of lineage-specific transcription factors (TFs) have emerged and facilitated the generation of highly pure populations of specific subtypes of neurons and glial cells in an easy, reproducible, and rapid manner. In this study, we review protocols based on this strategy to generate excitatory, inhibitory, dopaminergic, and motor neurons as well as astrocytes, oligodendrocytes, and microglia. In addition, we will discuss the main applications for cells generated with these protocols, including disease modeling, drug screening, and mechanistic studies. Finally, we will discuss the advantages and disadvantages of TF-based protocols and present our view of the future in this field.

Potential of Induced Pluripotent Stem Cells for Use in Gene Therapy: History, Molecular Bases, and Medical Perspectives.

Induced pluripotent stem cells (iPSCs) are defined as reprogrammed somatic cells exhibiting embryonic stem cell characteristics. Since their discovery in 2006, efforts have been made to utilize iPSCs in clinical settings. One of the promising fields of medicine, in which genetically patient-specific stem cells may prove themselves useful, is gene therapy. iPSCs technology holds potential in both creating models of genetic diseases and delivering therapeutic agents into the organism via auto-transplants, which reduces the risk of rejection compared to allotransplants. However, in order to safely administer genetically corrected stem cells into patients’ tissues, efforts must be made to establish stably pluripotent stem cells and reduce the risk of insertional tumorigenesis. In order to achieve this, optimal reprogramming factors and vectors must be considered. Therefore, in this review, the molecular bases of reprogramming safe iPSCs for clinical applications and recent attempts to translate iPSCs technology into the clinical setting are discussed.

CELLULAR AND MOLECULAR CHARACTERIZATION OF MESENCHYMAL STEM CELLS DERIVED FROM PLURIPOTENT STEM CELLS AND PROSPECTION OF APPLICATIONS IN REGENERATIVE MEDICINE

<h3>Background</h3> Stem cells have been studied for their properties on tissue regeneration and functional recovery of injured organs. Mesenchymal stem cells (MSCs) are prominent in this context as they promote the increase of activity and viability of several cell types through distinct mechanisms. However, biological samples of MSCs bring great variability in behavior and viability while in culture (e.g., cell population heterogeneity, senescence). Therefore, it is necessary to establish a standardized protocol for the generation of high-quality stable cell cultures. In this sense, the use of pluripotent stem cells (PSCs) in derivation protocols for generation of MSC-like cells (PSC-MSCs) is an alternative, since PSCs maintain their characteristics consistently under suitable culture conditions. <h3>Methods</h3> Pluripotent stem cells (HES3) grown as cell clumps were cultured in ultralow attachment plates in aggregation medium for 24 h under hypoxia to form embryoid bodies (EBs). Mesodermal differentiation was induced in the EBs during 4 days when CD56 mesodermal marker was evaluated by FACS. Induced EBs were then plated into gelatin coatings and cultured in standard MSC medium containing FBS. After 4 passages the generated cells were evaluated for classical positive and negative MSC markers as defined by ISCT. <h3>Results</h3> Mesoderm induction was observed in induced EBs by CD56 expression (78%). During culture in FBS-containing medium, EBs attached and migrating cells presented fibroblast-like morphology. With progression of culture PSC-MSCs acquired epithelial-like morphology and slowly retrieved fibroblast-like morphology until passage 4 (P4). FACS analysis in P4 identified positive MSC markers CD90/CD140b in the majority of PSC-MSCs, and CD73/CD13 present in about one third of the cells. CD105 expression was not detected. Negative MSC markers CD34/CD11b/CD45/CD19/CD31/HLA-DR were not expressed by PSC-MSCs as well. About 26% of the cells expressed CD90 and CD73 simultaneously. Half of the analyzed cells still expressed CD56, indicating PSC-MSC may be less mature than adult MSCs. <h3>Conclusion</h3> PSC-MSC derivation protocol via mesodermal induction is feasible and the generated cells expressed positive MSC markers and lacked the negative ones. PSC-MSC were expanded <i>in vitro</i> for at least four passages. Further investigations will analyze other minimal criteria of MSC in the derived cells as trilineage differentiation potential, besides transcriptomic and karyotyping analysis.

Recent progress and new challenges in modeling of human pluripotent stem cell-derived blood-brain barrier.

The blood-brain barrier (BBB) is a semipermeable unit that serves to vascularize the central nervous system (CNS) while tightly regulating the movement of molecules, ions, and cells between the blood and the brain. The BBB precisely controls brain homeostasis and protects the neural tissue from toxins and pathogens. The BBB is coordinated by a tight monolayer of brain microvascular endothelial cells, which is subsequently supported by mural cells, astrocytes, and surrounding neuronal cells that regulate the barrier function with a series of specialized properties. Dysfunction of barrier properties is an important pathological feature in the progression of various neurological diseases. In vitro BBB models recapitulating the physiological and diseased states are important tools to understand the pathological mechanism and to serve as a platform to screen potential drugs. Recent advances in this field have stemmed from the use of pluripotent stem cells (PSCs). Various cell types of the BBB such as brain microvascular endothelial cells (BMECs), pericytes, and astrocytes have been derived from PSCs and synergistically incorporated to model the complex BBB structure in vitro. In this review, we summarize the most recent protocols and techniques for the differentiation of major cell types of the BBB. We also discuss the progress of BBB modeling by using PSC-derived cells and perspectives on how to reproduce more natural BBBs in vitro.

Development of pluripotent stem cell-based human tenocytes.

Human pluripotent stem cells (PSCs) are used as a platform for therapeutic purposes such as cell transplantation therapy and drug discovery. Another motivation for studying PSCs is to understand human embryogenesis and development. All cell types that make up the body tissues develop through defined trajectories during embryogenesis. For example, paraxial mesoderm is considered to differentiate into several cell types including skeletal muscle cells, chondrocytes, osteocytes, dermal fibroblasts, and tenocytes. Tenocytes are fibroblast cells that constitute the tendon. The step-wise narrowing fate decisions of paraxial mesoderm in the embryo have been modeled in vitro using PSCs; however, deriving tenocytes from human-induced PSCs and their application in cell therapy have long been challenging. PSC-derived tenocytes can be used for a source of cell transplantation to treat a damaged or ruptured tendon due to injury, disorder, or aging. In this review, we discuss the latest research findings on the use of PSCs for studying the biology of tenocyte development and their application in therapeutic settings.

Roles of Reactive Oxygen Species in Cardiac Differentiation, Reprogramming, and Regenerative Therapies

Reactive oxygen species (ROS) have been implicated in mechanisms of heart development and regenerative therapies such as the use of pluripotent stem cells. The roles of ROS mediating cell fate are dependent on the intensity of stimuli, cellular context, and metabolic status. ROS mainly act through several targets (such as kinases and transcription factors) and have diverse roles in different stages of cardiac differentiation, proliferation, and maturation. Therefore, further detailed investigation and characterization of redox signaling will help the understanding of the molecular mechanisms of ROS during different cellular processes and enable the design of targeted strategies to foster cardiac regeneration and functional recovery. In this review, we focus on the roles of ROS in cardiac differentiation as well as transdifferentiation (direct reprogramming). The potential mechanisms are discussed in regard to ROS generation pathways and regulation of downstream targets. Further methodological optimization is required for translational research in order to robustly enhance the generation efficiency of cardiac myocytes through metabolic modulations. Additionally, we highlight the deleterious effect of the host's ROS on graft (donor) cells in a paracrine manner during stem cell-based implantation. This knowledge is important for the development of antioxidant strategies to enhance cell survival and engraftment of tissue engineering-based technologies. Thus, proper timing and level of ROS generation after a myocardial injury need to be tailored to ensure the maximal efficacy of regenerative therapies and avoid undesired damage.

Insights into stem cell therapy for diabetic retinopathy: a bibliometric and visual analysis.

Stem cells have been confirmed to be involved in the occurrence and development of diabetic retinopathy; however, the underlying mechanisms remain unclear. In this study, we used Citespace software to visually analyze 552 articles exploring the stem cell-based treatment of diabetic retinopathy over the past 20 years, which were included in the Web of Science Core Collection. We found the following: (1) a co-citation analysis of the references cited by all 552 articles indicated 15 clusters. In cluster #0, representing the stem cell field, some highly cited landmark studies emerged between 2009–2013. For example, endothelial progenitor cells and diabetic retinopathy gradually received the full attention of scholars, in terms of their relationship and therapeutic prospects. Some researchers also verified the potential of adipose-derived stem cells to differentiate into stable retinal perivascular cells, using a variety of animal models of retinal vascular disease. All of these achievements provided references for the subsequent stem cell research. (2) An analysis of popular keywords among the 552 articles revealed that, during the past 20 years, a relative increase in basic research articles examining stem cells and endothelial progenitor cells for the treatment of diabetic retinopathy was observed. The contents of these articles primarily involved the expression of vascular endothelial growth factor, vascular regeneration, oxidative stress, and inflammatory response. (3) A burst analysis of keywords used in the 552 articles indicated that genetic and cytological research regarding the promotion of angiogenesis was an issue of concern from 2001 to 2012, including several studies addressing the expression of various growth factor genes; from 2014 to 2020, mouse models of diabetic retinopathy were recognized as mature animal models, and the most recent research has focused on macular degeneration, macular edema, neurodegeneration, and inflammatory changes in diabetic animal models. (4) Globally, the current authoritative studies have focused on basic research towards the stem cell treatment of diabetic retinopathy. Existing clinical studies are of low quality and have insufficient evidence levels, and their findings have not yet been widely accepted in clinical practice. Major challenges during stem cell transplantation remain, including stem cell heterogeneity, cell delivery, and the effective homing of stem cells to damaged tissue. However, clinical trials examining potential stem cell-based treatments of diabetic retinopathy, including the use of pluripotent stem cells, retinal pigment epithelial cells, bone marrow mesenchymal stem cells, and endothelial progenitor cells, are currently ongoing, and high-quality clinical evidence is likely to appear in the future, to promote clinical transformation.

Induced pluripotent stem cell-derived natural killer cells gene-modified to express chimeric antigen receptor-targeting solid tumors.

The use of allogeneic, pluripotent stem cell-derived immune cells for cancer immunotherapy has been the subject of recent research, including clinical trials. The use of pluripotent stem cells as the source for allogeneic immune cells facilitates stringent quality control of the final product, regarding efficacy, safety, and producibility. In this review, we have described the characteristics of natural killer (NK) cells from multiple cell sources, including pluripotent stem cells, the chimeric antigen receptor (CAR)-modification method and strategy for these NK cells, and the current and planned clinical trials of CAR-modified induced pluripotent stem cell-derived NK cells.