Isolation,Culture and Comprehensive Characterization of Biological Properties of Human Urine-Derived Stem Cells

Mesenchymal stem cells (MSCs) represent an attractive source within the field of tissue engineering. However, their harvesting often requires invasive medical procedures. Urine-derived stem cells (UDSCs) display similar properties to MSCs, and their obtention and further processing is non-invasive for the donors as well as low cost. Here, we offer a comprehensive analysis of their biological properties. The goal of this study was to analyze their morphology, stemness, differentiation potential and cytokine profile. We have successfully isolated UDSCs from 25 urine samples. First colonies emerged up to 9 days after the initial seeding. Cell doubling time was 45 ± 0.24 SD, and when seeded at the density of 100 cells/cm², they formed 42 ± 6.5 SD colonies within 10 days. Morphological analyzes revealed that two different types of the cell populations have been present. The first type had a rice-grain shape and the second one was characterized by a polyhedral shape. In several cell cultures, dome-shaped cells were observed as well. All examined UDSCs expressed typical MSC-like surface markers, CD73, CD90 and CD105. Moreover, conditioned media from UDSCs were harvested, and cytokine profile has been evaluated showing a significantly higher secretory rate of IL-8, IL-6 and chemokines MCP-1 and GM-CSF. We have also successfully induced human UDSCs into chondrogenic, osteogenic and myogenic cell lineages. Our findings indicate that UDSCs might have immense potential in the regeneration of the damaged tissues.     

Validation of SV2A-Targeted PET Imaging for Noninvasive Assessment of Neuroendocrine Differentiation in Prostate Cancer

Neuroendocrine prostate cancer (NEPC) is an aggressive and lethal variant of prostate cancer (PCa), and it remains a diagnostic challenge. Herein we report our findings of using synaptic vesicle glycoprotein 2 isoform A (SV2A) as a promising marker for positron emission tomography (PET) imaging of neuroendocrine differentiation (NED). The bioinformatic analyses revealed an amplified SV2A gene expression in clinical samples of NEPC versus castration-resistant PCa with adenocarcinoma characteristics (CRPC-Adeno). Importantly, significantly upregulated SV2A protein levels were found in both NEPC cell lines and tumor tissues. PET imaging studies were carried out in NEPC xenograft models with ¹⁸F-SynVesT-1. Although ¹⁸F-SynVesT-1 is not a cancer imaging agent, it showed a significant uptake level in the SV2A⁺ tumor (NCI-H660: 0.70 ± 0.14 %ID/g at 50–60 min p.i.). The SV2A blockade resulted in a significant reduction of tumor uptake (0.25 ± 0.03 %ID/g, p = 0.025), indicating the desired SV2A imaging specificity. Moreover, the comparative PET imaging study showed that the DU145 tumors could be clearly visualized by ¹⁸F-SynVesT-1 but not ⁶⁸Ga-PSMA-11 nor ⁶⁸Ga-DOTATATE, further validating the role of SV2A-targeted imaging for noninvasive assessment of NED in PCa. In conclusion, we demonstrated that SV2A, highly expressed in NEPC, can serve as a promising target for noninvasive imaging evaluation of NED.     

Pulsed Electromagnetic Field Stimulation in Osteogenesis and Chondrogenesis: Signaling Pathways and Therapeutic Implications

Mesenchymal stem cells (MSCs) are the main cell players in tissue repair and thanks to their self-renewal and multi-lineage differentiation capabilities, they gained significant attention as cell source for tissue engineering (TE) approaches aimed at restoring bone and cartilage defects. Despite significant progress, their therapeutic application remains debated: the TE construct often fails to completely restore the biomechanical properties of the native tissue, leading to poor clinical outcomes in the long term. Pulsed electromagnetic fields (PEMFs) are currently used as a safe and non-invasive treatment to enhance bone healing and to provide joint protection. PEMFs enhance both osteogenic and chondrogenic differentiation of MSCs. Here, we provide extensive review of the signaling pathways modulated by PEMFs during MSCs osteogenic and chondrogenic differentiation. Particular attention has been given to the PEMF-mediated activation of the adenosine signaling and their regulation of the inflammatory response as key player in TE approaches. Overall, the application of PEMFs in tissue repair is foreseen: (1) in vitro: to improve the functional and mechanical properties of the engineered construct; (2) in vivo: (i) to favor graft integration, (ii) to control the local inflammatory response, and (iii) to foster tissue repair from both implanted and resident MSCs cells.     

Glutamatergic Neurons Differentiated from Embryonic Stem Cells: An Investigation of Differentiation and Associated Diseases

Neurons that have been derived from various types of stem cells have recently undergone significant study due to their potential for use in various aspects of biomedicine. In particular, glutamatergic neurons differentiated from embryonic stem cells (ESCs) potentially have many applications in both basic research and regenerative medicine. This review summarized the literatures published thus far and focused on two areas related to these applications. Firstly, these neurons can be used to investigate neuronal signal transduction during differentiation and this means that the genes/proteins/markers involved in this process can be identified. In this way, the dynamic spatial and temporal changes associated with neuronal morphology can be investigated relatively easily. Such an in vitro system can also be used to study how neurons during neurogenesis integrate into normal tissue. At the same time, the integration, regulation and functions of extracellular matrix secretion, various molecular interactions, various ion channels, the neuronal microenvironment, etc., can be easily traced. Secondly, the disease-related aspects of ESC-derived glutamatergic neurons can also be studied and then applied therapeutically. In the future, greater efforts are needed to explore how ESC-differentiated glutamatergic neurons can be used as a neuronal model for the study of Alzheimer’s disease (AD) mechanistically, to identify possible therapeutic strategies for treating AD, including tissue replacement, and to screen for drugs that can be used to treat AD patients. With all of the modern technology that is available, translational medicine should begin to benefit patients soon.     

The Emerging Regulatory Role of Circular RNAs in Periodontal Tissues and Cells

Periodontitis is a chronic complex inflammatory disease associated with a destructive host immune response to microbial dysbiosis, leading to irreversible loss of tooth-supporting tissues. Regeneration of functional periodontal soft (periodontal ligament and gingiva) and hard tissue components (cementum and alveolar bone) to replace lost tissues is the ultimate goal of periodontal treatment, but clinically predictable treatments are lacking. Similarly, the identification of biomarkers that can be used to accurately diagnose periodontitis activity is lacking. A relatively novel category of molecules found in oral tissue, circular RNAs (circRNAs) are single-stranded endogenous, long, non-coding RNA molecules, with covalently circular-closed structures without a 5’ cap and a 3’ tail via non-classic backsplicing. Emerging research indicates that circRNAs are tissue and disease-specific expressed and have crucial regulatory functions in various diseases. CircRNAs can function as microRNA or RNA binding sites or can regulate mRNA. In this review, we explore the biogenesis and function of circRNAs in the context of the emerging role of circRNAs in periodontitis pathogenesis and the differentiation of periodontal cells. CircMAP3K11, circCDK8, circCDR1as, circ_0062491, and circ_0095812 are associated with pathological periodontitis tissues. Furthermore, circRNAs are expressed in periodontal cells in a cell-specific manner. They can function as microRNA sponges and can form circRNA–miRNA–mRNA networks during osteogenic differentiation for periodontal-tissue (or dental pulp)-derived progenitor cells.     

High Glucose Exposure Impairs L-Cell Differentiation in Intestinal Organoids: Molecular Mechanisms and Clinical Implications

Intestinal organoids are used to analyze the differentiation of enteroendocrine cells (EECs) and to manipulate their density for treating type 2 diabetes. EEC differentiation is a continuous process tightly regulated in the gut by a complex regulatory network. However, the effect of chronic hyperglycemia, in the modulation of regulatory networks controlling identity and differentiation of EECs, has not been analyzed. This study aimed to investigate the effect of glucotoxicity on EEC differentiation in small intestinal organoid platforms. Mouse intestinal organoids were cultured in the presence/absence of high glucose concentrations (35 mM) for 48 h to mimic glucotoxicity. Chronic hyperglycemia impaired the expression of markers related to the differentiation of EEC progenitors (Ngn3) and L-cells (NeuroD1), and it also reduced the expression of Gcg and GLP-1 positive cell number. In addition, the expression of intestinal stem cell markers was reduced in organoids exposed to high glucose concentrations. Our data indicate that glucotoxicity impairs L-cell differentiation, which could be associated with decreased intestinal stem cell proliferative capacity. This study provides the identification of new targets involved in new molecular signaling mechanisms impaired by glucotoxicity that could be a useful tool for the treatment of type 2 diabetes.     

Anti-Inflammatory Fibronectin-AgNP for Regulation of Biological Performance and Endothelial Differentiation Ability of Mesenchymal Stem Cells

The engineering of vascular regeneration still involves barriers that need to be conquered. In the current study, a novel nanocomposite comprising of fibronectin (denoted as FN) and a small amount of silver nanoparticles (AgNP, ~15.1, ~30.2 or ~75.5 ppm) was developed and its biological function and biocompatibility in Wharton’s jelly-derived mesenchymal stem cells (MSCs) and rat models was investigated. The surface morphology as well as chemical composition for pure FN and the FN-AgNP nanocomposites incorporating various amounts of AgNP were firstly characterized by atomic force microscopy (AFM), UV-Visible spectroscopy (UV-Vis), and Fourier-transform infrared spectroscopy (FTIR). Among the nanocomposites, FN-AgNP with 30.2 ppm silver nanoparticles demonstrated the best biocompatibility as assessed through intracellular ROS production, proliferation of MSCs, and monocytes activation. The expression levels of pro-inflammatory cytokines, TNF-α, IL-1β, and IL-6, were also examined. FN-AgNP 30.2 ppm significantly inhibited pro-inflammatory cytokine expression compared to other materials, indicating superior performance of anti-immune response. Mechanistically, FN-AgNP 30.2 ppm significantly induced greater expression of vascular endothelial growth factor (VEGF) and stromal-cell derived factor-1 alpha (SDF-1α) and promoted the migration of MSCs through matrix metalloproteinase (MMP) signaling pathway. Besides, in vitro and in vivo studies indicated that FN-AgNP 30.2 ppm stimulated greater protein expressions of CD31 and von Willebrand Factor (vWF) as well as facilitated better endothelialization capacity than other materials. Furthermore, the histological tissue examination revealed the lowest capsule formation and collagen deposition in rat subcutaneous implantation of FN-AgNP 30.2 ppm. In conclusion, FN-AgNP nanocomposites may facilitate the migration and proliferation of MSCs, induce endothelial cell differentiation, and attenuate immune response. These finding also suggests that FN-AgNP may be a potential anti-inflammatory surface modification strategy for vascular biomaterials.     

Advances and Perspectives in Dental Pulp Stem Cell Based Neuroregeneration Therapies

Human dental pulp stem cells (hDPSCs) are some of the most promising stem cell types for regenerative therapies given their ability to grow in the absence of serum and their realistic possibility to be used in autologous grafts. In this review, we describe the particular advantages of hDPSCs for neuroregenerative cell therapies. We thoroughly discuss the knowledge about their embryonic origin and characteristics of their postnatal niche, as well as the current status of cell culture protocols to maximize their multilineage differentiation potential, highlighting some common issues when assessing neuronal differentiation fates of hDPSCs. We also review the recent progress on neuroprotective and immunomodulatory capacity of hDPSCs and their secreted extracellular vesicles, as well as their combination with scaffold materials to improve their functional integration on the injured central nervous system (CNS) and peripheral nervous system (PNS). Finally, we offer some perspectives on the current and possible future applications of hDPSCs in neuroregenerative cell therapies.