Tissue engineering and regenerative medicine最新文献

Background: Granulosa-like cells (GLCs) were differentiated from human endometrium-derived induced pluripotent stem cells (heiPSCs). This study aimed to establish a GLC differentiation protocol as a defined means to produce autologous estradiol (in vitro), highlighting its therapeutic potential as an alternative to conventional menopausal hormone therapy.

Methods: Endometrial cells from hysterectomy specimens were reprogrammed into iPSCs using episomal vectors encoding SOX2, OCT4, c-MYC, and KLF4. Differentiation was induced using Activin A and CHIR99021 for mesoderm induction, followed by BMP4, Follistatin, and bFGF. Gene expression, flow cytometry, and immunofluorescence were analyzed at each stage. Estradiol production was quantified by ELISA, and its effect on endometrial cell proliferation was evaluated by MTT assay.

Results: Results: The roles of small molecules and growth factors in directing stem cells toward functional chondrocytes are also discussed. Additionally, we briefly examine the emerging integration of artificial intelligence (AI) in cartilage tissue engineering. AI applications such as predicting differentiation outcomes, monitoring chondrogenic progression in real-time, and identifying small-molecule enhancers are poised to accelerate discovery and standardization in the field.

Conclusion: GLCs expressing the key markers and capable of E2 production were successfully derived from heiPSCs, which may be developed as a novel source for menopausal hormone therapy.

Background: Treatment for nonunion in long bones remains a clinical need. Collagen sponges loaded with recombinant human bone morphogenetic protein 2 (rhBMP-2) are potential grafts but have limited FDA-approved applications due to safety concerns associated with rapid collagen resorption and burst release of rhBMP-2 in vivo. This work investigates keratin proteins obtained from human hair as a potential rhBMP-2 biomaterial carrier. Keratins are an appealing carrier because the extent of disulfide crosslinking can be modulated by the form of keratin present, thus allowing control over the rate of scaffold degradation.

Methods: The two forms of keratin used to formulate carriers were reductively extracted keratin called kerateine (KTN) that can form disulfide crosslinks and oxidatively extracted keratin called keratose (KOS) that does not form disulfide crosslinks. Five formulations of freeze-dried keratin scaffolds containing variable amounts of KOS and KTN were fabricated and implanted into a critically-sized rat femur defect model.

Results: A 50:50 KOS:KTN formulation with rhBMP-2 showed the same level of bone bridging, bone mineral density, and bone volume as collagen with rhBMP-2 by 8 weeks as determined by μ-CT. Scaffolds with the 50:50 KOS:KTN or 100% KTN showed approximately fourfold higher retention of fluorescently-labeled rhBMP-2 at the implant site 1, 3, or 7 days post-implant compared to collagen or 100% KOS scaffolds. The increased retention with 50:50 KOS:KTN or 100% KTN correlated with decreased levels of fluorescent rhBMP-2 in distal organs.

Conclusions: Keratin scaffolds could provide comparable levels of bone regeneration as collagen carriers with improved safety profiles suitable for use in long bone nonunions.

Background: Sarcopenia, a musculoskeletal disease associated with aging or certain factors, is characterized by a reduction in muscle mass, strength, and performance. Dexamethasone (DEX)-induced muscular atrophy in animals, which shows a significant decrease in muscle mass, strength, and function, serves as a model for sarcopenia. Mesenchymal stem cell-based therapies, particularly those using 3D cultured spheroids, have emerged as a prominent area in muscle regeneration. Previous research has demonstrated that tonsil-derived mesenchymal stem cells (TMSCs) can differentiate into skeletal muscle cells (SKMCs) that exhibit attributes of skeletal muscles.

Methods: Spheroids formed from TMSC-derived skeletal muscle cells (TMSC-SKMC-spheroids) were produced using microwells and subsequently transplanted into a sarcopenia model. This model utilized a dexamethasone (DEX)-induced muscular atrophy rat to mimic sarcopenia. The effectiveness of TMSC-SKMC-spheroid transplantation was assessed through grip strength tests, running fatigue tests, measurements of gastrocnemius muscle thickness and weight, and histopathological evaluations.

Results: Post-transplantation, the rat models exhibited improvement in hind limb motor functions and gastrocnemius muscle regeneration. Additionally, the neuromuscular junctions in the gastrocnemius muscle of the transplantation group were restored.

Conclusion: These findings demonstrate the therapeutic potential of TMSC-SKMC-spheroids in the DEX-induced atrophy rat model and suggest their promise as a valuable therapeutic resource for sarcopenia caused by various factors.

Background: Damage to the peripheral nerve significantly impairs quality and way of life. Despite the slow self-regeneration of the peripheral nervous system and advances in surgery, complete recovery after nerve injury remains elusive especially when there is loss of substance. Human olfactory ecto-mesenchymal stem cells (OEMSC) have potential for the treatment of peripheral nerve injury through the secretion of bioactive factors, such as proteins, cytokines, chemokines as well as the release of extracellular vesicles (EV). The current research investigates the therapeutic effects of a venous bridge, filled with freshly purified or cryoconserved OEMSC-derived EVs after a peroneal nerve loss of substance.

Methods: A 7 mm defect of peroneal nerve was bridged with a vein into which freshly purified or cryoconserved OEMSC-derived EVs were injected or not. These groups were compared with unoperated rats (Control) and autografted rats with the nerve sutured in inverted position (Gold Standard). Three months after surgery, nerve repair was analyzed by measuring locomotor function, muscle mechanical properties, muscle mass, axon number, and myelination.

Results: We observed that OEMSC-derived EVs significantly (i) increased locomotor recovery, (ii) partially maintained the contractile phenotype of the target muscle, and (iii) augmented the number of growing axons. Freshly purified EVs exerted a better recovery than the Gold Standard.

Conclusion: These results demonstrate that EVs display a positive effect on peripheral nerve regeneration, similarly to what has been observed with OEMSCs, the cells from which they originate. They represent an alternative to cellular therapies for peripheral nerve repair.

Background: Infertility has a significant impact on women, affecting them both mentally and physically. Some of the current infertility-related diseases include intrauterine adhesions, endometriosis, polycystic ovary syndrome, primary ovarian insufficiency, and cancer. While existing treatments can help slow diseases progression and improve fertility for some patients, overall recovery rates remain low. The use of three-dimensional bioprinting (3D bioprinting) is becoming increasingly popular in clinical settings due to its high precision, customizable materials, and mechanical properties. In the state of infertility, its therapeutic potential is becoming more evident.

Methods: In this paper, we summarized the current treatment status of female infertility-related diseases, including cervical cancer, ovarian cancer, endometrial cancer, polycystic ovary syndrome, intrauterine adhesions, MRKH syndrome and other diseases, used databases such as PubMed, described the research progress and future development direction of 3D bioprinting in these diseases.

Results: 3D bioprinting technology could help repair damaged endometrial and ovarian tissue, and was able to create cell-loaded biological scaffolds to help restore the structure and function of affected organs. Furthermore, the development of organoids is opening new ways for research in regenerative medicine. It is expected that 3D bioprinting will not only be able to create organoid structures for research purposes but will also be utilized in clinical settings to effectively address infertility.

Conclusions: 3D bioprinting is gaining popularity in the clinical field due to its high resolution, adjustable composition, and mechanical qualities. Infertility-related disorders damage women by inflicting a psychological and functional double blow. According to the current research, the application of 3D bioprinting technology to help patients restore fertility function has endless possibilities in the future.

Background: The scaffold-free approach has emerged with a focus on creating cartilage-like tissues using cell pellets, cell spheroids, and cell sheets. However, complete repair of damaged cartilage using these tissues remains an ongoing challenge due to the limitation of thin structure and poor structural integrity.

Method: In this study, we proposed a novel method to produce scaffold-free cartilage by combining cell pellets and cell sheet technology as chondrocyte pellet-sheet tissues. The chondrocyte sheets acted as a support platform at the top and the bottom of the ten chondrocyte pellets. At day 7, the quality of the tissues cultured in a chondrogenic differentiation medium (CDM) and basal medium was compared using real-time PCR, immunofluorescence staining, proteomics, and atomic force microscopy (AFM).

Results: Our method supported the enhancement of tissue thickness. Compared to the control basal medium, the diameter and thickness of the chondrocyte pellet-sheet tissues in CDM were 1.47- and 2.21-fold increase, respectively. The level of mRNA expression and immunostaining of collagen type II were higher in the tissues cultured in CDM, compared to those in basal medium. Using proteomics, transferrin was found in both fresh and cultured CDM. The protein profiles of the tissues in CDM revealed the downregulation of actin and the upregulation of fibromodulin (FMOD), which related to the reorganization of cell shape and the production of cartilage ECM, respectively. Pathway analysis of chondrocyte pellet-sheet tissues in CDM also revealed the inhibition of RhoA and the presence of a TGFβ signaling pathway with SMAD protein signals. Moreover, Young's modulus indicating structural integrity of the tissues cultured in CDM (28.25 ± 13.13 kPa) was higher than those in basal medium (4.63 ± 2.25 kPa).

Conclusion: Combining chondrocyte pellets and sheets in CDM allowed the generation of thick tissues and enhanced structural integrity. The compacted structure of the tissues in CDM might inhibit actin expression via RhoA inhibition. Growth factors in CDM, especially transferrin might be involved in chondrogenic differentiation via TGFβ signaling pathway with SMAD protein signals.

Background: Sjögren's syndrome (SS) is a chronic autoimmune disease delineated by excessive lymphocyte infiltration to the lacrimal or salivary glands, leading to dry eye and dry mouth. Exosomes secreted from mesenchymal stem cells (MSC) are known to have anti-inflammatory and tissue regeneration abilities. This study endeavored to demonstrate the effect of MSC-derived exosomes on the clinical parameter of dry eyes and associated pathology in SS mouse model.

Methods: Exosomes obtained from bone marrow-derived human MSC (Catholic MASTER Cells) were injected into the subconjunctival sac of 17 weeks-old NOD/LtJ female mice once and sacrificed after 7 days, or administered topically as an eyedrop every day for 14 days, then sacrificed. Clinical dry eye parameters, including tear volume and corneal staining scores, density of goblet cells in the conjunctiva, pro-inflammatory cytokine expressions of cornea and conjunctiva, and the lacrimal glands were evaluated. Infiltration of inflammatory foci, and expression of B and T cells in the lacrimal glands were examined.

Results: Tear volume, corneal stain scores and density of goblet cells in conjunctiva were improved in the exosome-treated groups compared to the control group. Pro-inflammatory cytokine expressions were also reduced in the cornea and conjunctiva of the exosome-treated group. In the lacrimal glands of the exosome-treated mice, inflammatory foci infiltration and B cell marker expressions were significantly decreased.

Conclusions: Thus, this study demonstrated the amelioration of dry eyes with the administration of exosomes in SS animal model, suggesting promising therapeutic potential of MSC-derived exosomes in SS dry eyes.

Background: Neurotraumatic conditions, such as spinal cord injury, brain injury, and neurodegenerative conditions, such as amyotrophic lateral sclerosis, pose a challenge to the field of rehabilitation for its complexity and nuances in management. For decades, the use of cell therapy in treatment of neurorehabilitation conditions have been explored to complement the current, mainstay treatment options; however, a consensus for standardization of the cell therapy and its efficacy has not been reached in the medical community. This study aims to provide a comparative review on the very topic of cell therapy use in neurorehabilitation conditions in an attempt to bridge the gap in knowledge.

Methods: Studies were searched from the PubMed database published from 2014 to 2024 employing the terms including but not exclusive to "spinal cord injury," "brain injury," "amyotrophic lateral sclerosis," "regenerative medicine," "cell therapy," and "stem cell." Following the PRISMA 2020 statement, the studies were screened, included, and excluded. Thirty three studies were identified and selected for this review.

Results: Countless researchers investigated the efficacy of various stem cell products for the treatment of numerous neurotraumatic conditions, such as spinal cord injury, traumatic brain injury, and neurodegenerative conditions such as amyotrophic lateral sclerosis. The recent decade of studies suggest that in neurotraumatic conditions, bone-marrow-derived and neural stem cells can be effective, and in neurodegenerative conditions, such as ALS, mesenchymal and neural stem cells can be efficacious.

Conclusion: Emerging data from the latest research is encouraging to the patients suffering from neurotraumatic and neurodegenerative conditions, which present themselves as a need for further studies with improved standardization in study design, including cell source specification, differentiation and culture method, and outcome measures to ensure a wide applicability.

Background: Mycophenolic acid (MPA) is a well-known inhibitor of inosine monophosphate dehydrogenases (IMPDH), and our previous study also highlighted its inhibitory effects on histone deacetylases (HDACs). This study aims to evaluate MPA as a potential regulator of cell differentiation and to explore its mechanisms as an inhibitor of IMPDHs and HDACs using tonsil-derived mesenchymal stem cells (TMSCs).

Methods: TMSCs were treated with MPA at various concentrations for 48 h to evaluate its cytotoxic effects. The MPA treatment conditions were then optimized to induce differentiation of TMSCs. To investigate the underlying mechanisms, protein levels of IMPDH, HDAC3 and the histone acetyltransferase p300 (EP300) were assessed.

Results: MPA even at the highest concentration (160 μM) showed strong cytostatic effects on TMSCs without inducing cell death. Pre-treatment with 10 μM MPA for 48 h promoted osteogenic differentiation while inhibiting adipogenic differentiation. IMPDH inhibition via siRNA mimicked these effects, increasing calcium mineralization in the differentiated TMSCs. Co-treatment with guanosine reversed these effects, highlighting guanine depletion as a key mechanism of inducing osteogenesis. MPA also reduced the protein levels of HDAC3 and histone acetyltransferase p300 (EP300), which were also reversed by guanosine supplementation. The knockdown of HDAC3 or EP300 using siRNAs enhanced osteogenesis, with guanosine reversing these effects. HDAC3 overexpression reversed the effects of MPA, decreasing calcium mineralization.

Conclusion: These findings suggest that MPA induces osteogenesis through guanine depletion, and that subsequent inhibition of HDAC3 and EP300 could be potential strategies for treating bone diseases.

Background: Recently, electrospun nanofiber composite scaffolds encaged with bioactive agents have gained prominence as an innovative therapy for managing full-thickness infectious wounds.

Methods: This study mainly focuses on the development and comprehensive characterization of multi-component polyvinyl alcohol (PVA)-based nanofiber scaffolds incorporating pupae oil (PO) and Prussian blue nanoparticles (PBNPs) using electrospinning technique for accelerated full thickness infectious wound healing.

Results: Scanning electron microscopy (SEM) photographs revealed a porous, interconnected fibrous structure with diameters ranging between 200 and 300 nm. Fourier-transform infrared spectroscopy confirmed the chemical compatibility and successful incorporation of PO and PBNPs into the scaffolds. The scaffolds exhibited optimal biodegradation over a two-week period and demonstrated appropriate water uptake capacity to absorb wound exudates. Furthermore, they displayed potent antibacterial and antibiofilm efficacy against Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa), as well as minimal microbial penetration across nanofiber scaffolds. In vitro studies on L-929 fibroblast cells indicated improved cell viability, migration, cell adhesion, and proliferation. In vivo evaluation in an infected rat model demonstrated rapid wound closure and improved tissue regeneration. Moreover, haematoxylin and eosin (H&E) and masson-trichome staining corroborated the scaffolds' excellent wound healing efficacy. Additionally, enzyme-linked immunosorbent assay demonstrated significant downregulation of key pro-inflammatory markers.

Conclusion: These results suggest that the bioinspired, multi-component PVA-based nanofiber scaffolds loaded with natural bioactive agents (PO and PBNPs), hold great potential as a therapeutic strategy for promoting enhanced healing of full-thickness infected wounds.