Figure 3 - available via license: Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International
Content may be subject to copyright.
Cell proliferation. Cell proliferation rate was significantly reduced in the iPSC-MSC-DLX3 group compared with the rate observed intheiPSC-MSC-GFP group on days 5 and 7. * P<0.05 vs. iPSC-MSC-DLX3. iPSC-MSC, induced pluripotent stem cell-derived mesenchymal stem cell; DLX3, distal-less homeobox 3.
Source publication
Osteoporosis is a disease characterized by the degeneration of bone structure and decreased bone mass. Induced pluripotent stem cell‑derived mesenchymal stem cells (iPSC‑MSCs) have multiple advantages that make them ideal seed cells for bone regeneration, including high‑level proliferation, multi‑differentiation potential and favorable immune compa...
Context in source publication
Context 1
... was 1.00±0.05 and 5.84±0.89, respectively (Fig. 2B). Moreover, the western blotting results demonstrated that the relative DLX3 expression in iPSC-MSC-GFP and iPSC-MSC-DLX3 was 1.00±0.10 and 4.05±0.81, respectively (Fig. 2C and D). Based on these results, it was suggested that the DLX3 gene was successfully transfected into iPSC-MSCs. (Fig. 3). There was no significant difference in cell numbers between the two groups on days 1 and 3 (P>0.05). However, iPSC-MSC-DLX3 demonstrated significantly lower proliferative activity compared with the iPSC-MSC-GFP group on days 5 and 7 ...
Citations
... DLX3 mRNA is expressed at relatively high levels in osteoblasts and stimulates osteoblastic differentiation [32]. DLX3 also influences cell proliferation and enhances the osteogenic differentiation of iPSC-MSCs [33]. Deletion of DLX3 in neural crest cells is associated with decreased bone formation and mineralization in craniofacial bones [34]. ...
Dental follicle cells (DFCs) promote bone regeneration in vivo and in vitro. Circular RNAs (circRNAs) play crucial roles in bone development and regeneration. Our previous study demonstrated the upregulation of circFgfr2 expression during the osteogenic differentiation of DFCs. However, the molecular mechanisms and functional roles of circFgfr2 in DFCs osteogenesis remain unclear. In this study, we aimed to investigate the subcellular localization of circFgfr2 in DFCs using fluorescence in situ hybridization. In vitro investigations demonstrated that circFgfr2 overexpression promoted osteogenic differentiation, as evidenced by real-time quantitative polymerase chain reaction. By integrating the outcomes of bioinformatics analyses, dual luciferase reporter experiments, and chromatin isolation by RNA purification, we identified circFgfr2 as a sponge for miR-133a-3p, a key regulator of osteogenic differentiation. Moreover, miR-133a-3p suppressed osteogenic differentiation by targeting DLX3 and RUNX2 in DFCs. We validated that circFgfr2 promoted the osteogenic differentiation of DFCs through the miR-133a-3p/DLX3 axis. To further investigate the therapeutic potential of circFgfr2 in bone regeneration, we conducted in vivo experiments and histological analyses. Overall, these results confirmed the crucial role of circFgfr2 in promoting osteogenesis. In summary, our findings demonstrated that the circFgfr2/miR-133a-3p/DLX3 pathway acts as a cascade, thereby identifying circFgfr2 as a promising molecular target for bone tissue engineering.
... DLX3 exerts a substantial effect on embryogenesis and organ development, such as epidermis and ectodermal appendages (44,45). DLX3 promoted osteogenic differentiation of BMSCs and the induced pluripotent stem cell-derived mesenchymal stem cells (46,47). DLX3 overexpression promoted the differentiation of BMSCs into osteoblasts through the Wnt/beta-catenin signaling-mediated histone methylation of DKK4 (26). ...
Osteoporosis is a complex multifactorial disorder linked to various risk factors and medical conditions. Bone marrow-derived mesenchymal stem cell (BMSC) dysfunction potentially plays a critical role in osteoporosis pathogenesis. Herein, the study identified that miR-4739 was upregulated in BMSC cultures harvested from osteoporotic subjects. BMSCs were isolated from normal and osteoporotic bone marrow tissues and identified for their osteogenic differentiation potential. In osteoporotic BMSCs, miR-4739 overexpression significantly inhibited cell viability, osteoblast differentiation, mineralized nodule formation, and heterotopic bone formation, whereas miR-4739 inhibition exerted opposite effects. Through direct binding, miR-4739 inhibited distal-less homeobox 3 (DLX3) expression. In osteoporotic BMSCs, DLX3 knockdown also inhibited BMSC viability and osteogenic differentiation. Moreover, DLX3 knockdown partially attenuated the effects of miR-4739 inhibition upon BMSCs. Altogether, the miR-4739/DLX3 axis modulates the capacity of BMSCs to differentiate into osteoblasts, which potentially plays a role in osteoporosis pathogenesis. The in vivo and clinical functions of the miR-4739/DLX3 axis require further investigation.
... Besides that, Alp staining and mineralized nodule counting revealed a significantly higher number of mineralized nodules in iPSC-MSC-DLX3 over the control group. Taken together, this study demonstrated the positive effect of DLX3 on osteogenic differentiation; however, the exact mechanisms of its action have not been fully understood yet [83]. Another recently found enhancer of osteogenic differentiation is menaquinone-7 (MK-7), in which a positive effect was proven on iPSC-MSCs, exhibiting a notable increase in Runx-2 expression, Alp activity, and collagen deposition [84]. ...
Oral and craniofacial bone defects caused by congenital disease or trauma are widespread. In the case of severe alveolar bone defect, autologous bone grafting has been considered a “gold standard”; however, the procedure has several disadvantages, including limited supply, resorption, donor site morbidity, deformity, infection, and bone graft rejection. In the last few decades, bone tissue engineering combined with stem cell-based therapy may represent a possible alternative to current bone augmentation techniques. The number of studies investigating different cell-based bone tissue engineering methods to reconstruct alveolar bone damage is rapidly rising. As an interdisciplinary field, bone tissue engineering combines the use of osteogenic cells (stem cells/progenitor cells), bioactive molecules, and biocompatible scaffolds, whereas stem cells play a pivotal role. Therefore, our work highlights the osteogenic potential of various dental tissue-derived stem cells and induced pluripotent stem cells (iPSCs), the progress in differentiation techniques of iPSCs into osteoprogenitor cells, and the efforts that have been made to fabricate the most suitable and biocompatible scaffold material with osteoinductive properties for successful bone graft generation. Moreover, we discuss the application of stem cell-derived exosomes as a compelling new form of “stem-cell free” therapy.
... Several studies have shown that iPSC-derived cells have effectively promoted bone repair and angiogenesis. [44][45][46][47] Human iPSCderived osteoblasts and osteoclasts were co-cultured with macrophages on hydroxyapatite-coated poly lacticco-glycolic acid/poly L-lactic acid scaffold. 16 Subsequent in vivo implantation of co-cultured osteoblasts and osteoclasts in rodents showed a mature bone-like growth. ...
The process of bone repair has always been a natural mystery. Although bones do repair themselves, supplemental treatment is required for the initiation of the self-regeneration process. Predominantly, surgical procedures are employed for bone regeneration. Recently, cell-based therapy for bone regeneration has proven to be more effective than traditional methods, as it eliminates the immune risk and painful surgeries. In clinical trials, various stem cells, especially mesenchymal stem cells, have shown to be more efficient for the treatment of several bone-related diseases, such as non-union fracture, osteogenesis imperfecta, osteosarcoma, and osteoporosis. Furthermore, the stem cells grown in a suitable three-dimensional scaffold support were found to be more efficient for osteogenesis. It has been shown that the three-dimensional bioscaffolds support and simulate an in vivo environment, which helps in differentiation of stem cells into bone cells. Bone regeneration in patients with bone disorders can be improved through modification of stem cells with several osteogenic factors or using stem cells as carriers for osteogenic factors. In this review, we focused on the various types of stem cells and scaffolds that are being used for bone regeneration. In addition, the molecular mechanisms of various transcription factors, signaling pathways that support bone regeneration and the senescence of the stem cells, which limits bone regeneration, have been discussed.
... Osteoporosis (OP), a systemic bone metabolic disease, is characterized by decreased bone mass, decreased bone density, and degeneration of bone microstructure [1]. OP tends to occur in the elderly [2]. ...
Osteoporosis (OP) is a systemic bone metabolic disease. Promotion of osteoblast proliferation and inhibition of cell apoptosis may be helpful for the prevention and clinical treatment of OP. In the current study, we focused on the expression changes and clinical values of lncRNA ROR and miR-145-5p in OP clinical serum samples, and investigated the interactive modulation effect of ROR/miR-145-5p on osteoblast function. Serum samples were obtained from 82 OP patients and 79 healthy individuals. MC3T3-E1 was applied for the cell experiments. Levels of lncRNA ROR and miR-145-5p were detected using qRT-PCR. Transient transfection was performed to regulate gene levels in cells, and cell proliferation and apoptosis were detected. A reciprocal correlation between lncRNA ROR and miR-145-5p was explored. LncRNA ROR was downregulated, and miR-145-5p was overexpressed in OP patients. The combined diagnosis of ROR and miR-145-5p showed good diagnostic value for OP. ROR knockdown promoted the MC3T3-E1 cell apoptosis and inhibited cell proliferation. Luciferase reporting assay verified the target relationship between ROR and miR-145-5p. MiR-145-5p downregulation reversed ROR silence mediated effect on MC3T3-E1 cell proliferation and apoptosis. LncRNA ROR is downregulated and miR-145-5p is highly expressed in OP patients. ROR knockdown may inhibit osteoblast proliferation via targeting miR-145-5p. It may provide a theoretical basis and experimental basis for ROR to be a potential target for the treatment of OP.
Background
Mesenchymal stem cells (MSCs) possess the ability to self-replicate, self-proliferate, and differentiate into various cell types, and hence have been extensively studied in the field of regenerative medicine. Despite the promising clinical applications of MSCs, their limited quantity and in vitro expansion potential from human tissues remain major concerns. Alternatively, MSCs can be derived from human embryonic stem cells (hESCs) that share similar phenotypic features, making hESC-MSCs a potential candidate for cell therapy. Our study aimed to investigate the efficacy of 3D-ESC-MSCs, obtained through a 3D differentiation system, as an immunoregulatory agent for treating liver damage caused by acetaminophen (APAP).
Methods
We differentiated human ESCs into MSCs using a 3D culture method involving a horizontal shaker. We characterized MSCs by detecting surface-specific markers through flow cytometry and qPCR, and validated their differentiation potential using in vitro lipid, bone, and cartilage differentiation assays. MSC proliferation and safety were tested using MTT, cell survival at 4°C, and nude mice tumorigenicity assays. The immune regulatory potential of 3D-ESC-MSCs was studied by transfecting polyI:C into these cells. We further investigated the effects of 3D-ESC-MSCs on APAP-induced liver injury by preconditioning hepatocyte cell line L-O2 with 3D-ESC-MSC conditioned medium and evaluating their cell viability through MTT assay. Additionally, we assessed the number of surviving cells following co-culturing with L-O2 cells stimulated with APAP. Finally, we administered 3D-ESC-MSCs to mice, via tail vein injection, with APAP-induced acute liver injury, and analyzed the repair effects by detecting ALT and AST levels in mouse serum, creating liver pathological sections, and HE staining.
Results
3D-ESC-MSCs were positive for CD73, CD90, and CD105 surface markers, and negative for hematopoietic markers CD45 and HLA-DR in. The cells expressed low levels of pluripotent genes OCT4 and NANOG. Compare to umbilical cord mesenchymal stem cells (UCMSCs), 3D-ESC-MSCs displayed excellent proliferation and low-temperature resistance, and lower concentrations of polyI:C were required to induce immune regulatory genes IDO1, IF71, IRF7, and ISG15. They also exhibited higher expression levels of immunomodulatory。In vitro experiments demonstrated that the conditioned medium of 3D-ESC-MSCs increased L-O2 cell activity under low concentrations of APAP, and the survival of L-O2 cells co-cultured with 3D-ESC-MSCs was higher compared to L-02 cells cultured alone under the same conditions. Animal experiments revealed that the ALT and AST levels in APAP-treated mice injected with 3D-ESC-MSCs were reduced, and the necrotic area of the liver in the 3D-ESC-MSC group was reduced. The therapeutic effect was similar to that of the UCMSC group.
Conclusions
3D-ESC-MSCs, differentiated from ESCs, exhibit stronger immunomodulatory effect and can be utilized to repair acute liver injury caused by APAP. This study highlights the clinical potential of 3D-ESC-MSCs in treating human diseases.
The foundations of cell reprogramming were laid by Yamanaka and co-workers, who showed that somatic cells can be reprogrammed into pluripotent cells (induced pluripotency). Since this discovery, the field of regenerative medicine has seen advancements. For example, because they can differentiate into multiple cell types, pluripotent stem cells are considered vital components in regenerative medicine aimed at the functional restoration of damaged tissue. Despite years of research, both replacement and restoration of failed organs/tissues have remained elusive scientific feats. However, with the inception of cell engineering and nuclear reprogramming, useful solutions have been identified to counter the need for compatible and sustainable organs. By combining the science underlying genetic engineering and nuclear reprogramming with regenerative medicine, scientists have engineered cells to make gene and stem cell therapies applicable and effective. These approaches have enabled the targeting of various pathways to reprogramme cells, i.e., make them behave in beneficial ways in a patient-specific manner. Technological advancements have clearly supported the concept and realization of regenerative medicine. Genetic engineering is used for tissue engineering and nuclear reprogramming and has led to advances in regenerative medicine. Targeted therapies and replacement of traumatized, damaged, or aged organs can be realized through genetic engineering. Furthermore, the success of these therapies has been validated through thousands of clinical trials. Scientists are currently evaluating induced tissue-specific stem cells (iTSCs), which may lead to tumour-free applications of pluripotency induction. In this review, we present state-of-the-art genetic engineering that has been used in regenerative medicine. We also focus on ways that genetic engineering and nuclear reprogramming have transformed regenerative medicine and have become unique therapeutic niches.
Mesenchymal stem cells (MSCs) represent the most clinically used stem cells in regenerative medicine. However, due to the disadvantages with primary MSCs, such as limited cell proliferative capacity and rarity in the tissues leading to limited MSCs, gradual loss of differentiation during in vitro expansion reducing the efficacy of MSC application, and variation among donors increasing the uncertainty of MSC efficacy, the clinical application of MSCs has been greatly hampered. MSCs derived from human pluripotent stem cells (hPSC-MSCs) can circumvent these problems associated with primary MSCs. Due to the infinite self-renewal of hPSCs and their differentiation potential towards MSCs, hPSC-MSCs are emerging as an attractive alternative for regenerative medicine. This review summarizes the progress on derivation of MSCs from human pluripotent stem cells, disease modelling and drug screening using hPSC-MSCs, and various applications of hPSC-MSCs in regenerative medicine. In the end, the challenges and concerns with hPSC-MSC applications are also discussed.
