Search Results (173)

The temporomandibular joint (TMJ) relies on specialized progenitor cells for tissue maintenance and repair. We characterized TMJ-derived progenitor cells in mice and investigated the role of Evc2-mediated Hedgehog signaling. Progenitor cells from the anterior TMJ exhibited greater colony-forming capacity and an elongated morphology, while posterior cells were cuboidal, highlighting regional heterogeneity. TMJ-derived progenitors demonstrated multipotency, differentiating into osteogenic and chondrogenic lineages. Gli1-expressing, slow-cycling cells localized to the ligament attachment regions, initially accumulating there and not overlapping with specialized cells (Col1⁺ cells). Conditional Evc2 disruption in Gli1-expressing cells paradoxically augmented expression of Gli1 and mechanosensors (Yap, Wwtr1, Piezo1), and produced more confluent, rapidly expanding colonies. We hypothesize that these colonies are primarily composed of transit amplifying cells (TACs), which may proliferate robustly but face challenges in terminal differentiation. These results reveal critical roles for EVC2 and regional progenitor cell diversity in TMJ regenerative biology and suggest that targeting cell signaling and mechanical factors may inform novel strategies for TMJ disorder therapies. Full article

Background: Adipose-derived mesenchymal stem/stromal cells (ADSCs) are gaining recognition in regenerative medicine for their potential for adipogenic, osteogenic, and chondrogenic differentiation, as well as their immunomodulatory properties. However, ADSC-based therapies focus either on differentiation for tissue replacement or on counteracting unrestrained inflammation to prevent tissue destruction and initiate regeneration. Here, we aim to examine the immunomodulatory potential of osteogenically differentiated ADSCs by analyzing their proteomic profile. Methods: Using LC-MS/MS, we generated the proteomic profiles of differentiated and undifferentiated ADSCs and compared them with the Reactome database. Transcriptomic analysis was also performed and compared with the proteomic profile. Results: Comparison of the proteomic (499 up-regulated; 355 down-regulated) and transcriptomic (212 up-regulated; 232 down-regulated) profiles showed 60.1% concordance—both proteins and transcripts showed the same trend. Significantly upregulated proteins in differentiating ADSCs (−log₁₀ p > 5 and >10) were grouped into four categories: propensity for osteogenic differentiation; immunomodulation/immune/inflammatory response; cell senescence; and cell cycle regulation. Among those proteins, thirteen were reported to play roles in processes such as immunomodulation, inflammatory signaling, or transplant rejection. Conclusions: We observed that differentiating ADSCs might still exert immunomodulatory effects, which could be used in the treatment of, e.g., bone defects. Full article

Large-scale expansion of human mesenchymal stem cells (hMSCs) remains a major challenge due to the intrinsic trade-off between cell proliferation and the maintenance of multipotency in conventional culture systems. Stiff substrates, such as tissue culture polystyrene or rigid hydrogels, promote rapid proliferation but induce progressive loss of stemness, whereas very soft matrices preserve multipotency at the expense of cell growth. To overcome this limitation, we developed mechanically soft, phase-separated gelatin–phenol/hyaluronic acid–phenol (Gtn-Ph/HA-Ph) hydrogels with precisely controlled microstructures via enzyme-mediated crosslinking. These hydrogels consist of HA-rich, dot-like domains embedded within a continuous Gtn-rich network, allowing for independent tuning of stiffness and domain architecture. On single-component Gtn-Ph hydrogels, hMSC proliferation increased with substrate stiffness, whereas soft hydrogels with a storage modulus (G′) of approximately 0.6 kPa markedly suppressed proliferation while preserving stemness marker expression, confirming the stiffness-dependent trade-off. In contrast, phase-separated Gtn-Ph/HA-Ph hydrogels supported robust hMSC proliferation even under soft mechanical conditions while maintaining high expression of stemness-associated markers. During long-term culture, hMSCs achieved a 68- to 195-fold increase in cumulative cell yield on soft Gtn-Ph/HA-Ph hydrogels (G′ = 0.5 kPa) compared with tissue culture polystyrene. Expression of α-smooth muscle actin (α-SMA) mRNA, encoded by the ACTA2 gene and associated with cellular senescence and fibrotic activation, was completely suppressed, while hMSCs retained robust adipogenic, osteogenic, and chondrogenic differentiation capacities. These results demonstrate that phase-separated Gtn-Ph/HA-Ph hydrogels effectively resolve the proliferation–multipotency dilemma in hMSC expansion and provide a promising platform for scalable manufacturing of therapeutic stem cells. Full article

Stem cell heterogeneity represents a critical yet underexplored variable in laser-assisted regenerative strategies. While photobiomodulation has been shown to influence mesenchymal stem cell (MSC) behavior, it remains unclear whether stem cell maturation status modulates responsiveness to Er,Cr:YSGG irradiation. This study investigated the differential response of magnetically separated STRO-1⁺ and STRO-1⁻ SHED subpopulations to low-power Er,Cr:YSGG laser stimulation (0.10 W and 0.25 W), focusing on viability, extracellular matrix production, and lineage commitment. STRO-1⁺ cells comprised 13.4% ± 1.2% of the total Stem Cells from Human Exfoliated Deciduous teeth (SHED) population. Laser exposure did not impair metabolic activity in either subpopulation. Collagen synthesis demonstrated a power- and time-dependent increase, with maximal enhancement observed in STRO-1⁺ cells at 0.25 W after 7 days. Laser irradiation selectively promoted osteogenic differentiation, as evidenced by increased alkaline phosphatase (ALP) expression at 0.10 W and enhanced mineral deposition, while chondrogenic potential remained unaffected and adipogenesis was reduced following 0.10 W exposure. These findings suggest that ALP expression is temporally and power-dependently modulated during osteogenic progression. Overall, Er,Cr:YSGG photobiomodulation does not uniformly affect heterogeneous SHED populations but modulates lineage allocation and extracellular matrix deposition in a maturation- and power-dependent manner. Integrating stem cell subpopulation selection with laser-based bioactivation may represent a strategy to refine regenerative endodontic and biomaterial-guided therapies. Full article

Articular cartilage, a highly specialised and avascular tissue, exhibits limited regenerative potential following trauma or degenerative conditions such as osteoarthritis (OA). Conventional surgical interventions, including microfracture and autologous chondrocyte implantation (ACI), have shown limited long-term efficacy due to donor site morbidity and restricted cell proliferation. In this context, mesenchymal stromal cells (MSCs) have emerged as a promising alternative owing to their multipotency, self-renewal capacity, and low immunogenicity. While bone marrow (BM) remains the traditional source of MSCs, recent studies have reported that peripheral blood-derived mesenchymal stromal cells (PB-MSCs) may possess chondrogenic, osteogenic, and adipogenic potential comparable to that of BM-derived MSCs. PB-MSCs can be harvested through minimally invasive methods, thereby avoiding the complications associated with BM aspiration. Experimental evidence indicates that PB-MSCs exhibit strong cell viability, proliferative potential, and the ability to synthesise cartilage-specific extracellular matrix proteins, such as type II collagen and sulphated glycosaminoglycans, within three-dimensional scaffolds. Immunophenotypically, PB-MSCs express mesenchymal markers including CD29, CD44, CD90, and CD105 while lacking hematopoietic markers CD34 and CD45. Flow cytometry analyses reveal that CD105⁺ populations increase following cryopreservation, highlighting their clinical utility. In contrast to these experimentally defined PB-MSCs, the term peripheral blood stem cells (PBSCs) is used in clinical studies to describe heterogeneous, non-cultured peripheral blood-derived cell preparations, typically enriched in hematopoietic stem and progenitor cells following granulocyte colony-stimulating factor (G-CSF) mobilisation, without full mesenchymal characterisation. In vitro studies confirm successful tri-lineage differentiation, whereas in vivo investigations have demonstrated effective cartilage regeneration using PB-based clinical approaches, including postoperative intra-articular administration of hyaluronic acid (HA) combined with PBSCs, as well as implantation of PBSCs covered with a collagen membrane. Furthermore, advancements in biomaterial engineering, such as poly(ethylene glycol)–cysteine–arginine–glycine–aspartic acid (PEG-CRGD) hydrogels, have enhanced PB-MSC adhesion, proliferation, and chondrogenic differentiation while promoting immunomodulation through M2 macrophage polarisation. Despite these promising outcomes, the available evidence remains limited and heterogeneous, with substantial variability in cell definitions, experimental models, and clinical study designs, which currently constrains definitive conclusions regarding clinical efficacy. Future research should focus on optimising isolation protocols, understanding molecular pathways governing PB-MSC chondrogenesis, and standardising clinical applications. Overall, PB-MSCs represent a viable, less invasive, and translationally relevant cell source for cartilage regeneration and regenerative orthopaedic therapies Full article

The achievements of regenerative medicine are based on methods of controlling stem cell division and differentiation. Electromagnetic fields stimulate cell differentiation by means of affecting calcium channels and cellular signaling. However, only a small part of the mechanisms underlying electromagnetic field effect on cells has been studied. The prospect of their use in tissue engineering as an addition or alternative to biochemical effects becomes clear in the course of numerous experiments. Electromagnetic stimulation enhances the effect of biochemical differentiation inducers and can cause the secretion of exosomes of special properties, which may serve as a therapeutic tool. For example, it has been shown that EMFs at 15 Hz and 2 mT increased the expression of chondrogenic differentiation markers SOX9 and COL2 in human bone-marrow MSCs by up to 3-fold (based on Parate et al.). Optimizing EMF parameters (e.g., 15–50 Hz, 1–2 mT) for specific cells and pathologies remains a key challenge of the studies in the field of tissue engineering. This review describes the electromagnetic field effect on the chondrogenic and osteogenic differentiation of MSCs of various origins, which is important for the musculoskeletal tissue recovery, as well as on inflammatory diseases in model animals. Full article

Biomorphic hydroxyapatite scaffolds derived from rattan wood (GreenBone) show significant promise in bone tissue engineering due to their inherent structural similarity to natural bone. Laser-drilled GreenBone scaffolds were studied for enhanced porosity, nutrient diffusion, cellular infiltration, and vascularisation. Patient-derived bone marrow mesenchymal stromal/stem cells (BMMSCs) and culture-expanded mesenchymal stem cells (cMSCs) demonstrated high cell viability (>90%), considerable adhesion, and extensive cytoskeletal organisation. Trilineage differentiation confirmed the multipotency of BMMSCs, with osteogenic, adipogenic, and chondrogenic markers being successfully expressed. BMMSCs and cMSCs exhibited enhanced differentiation and gene expression profiles. At week 4, key osteogenic and angiogenic genes such as BMP2, VEGFC, RUNX2, and COL1A1 showed elevated expression, indicating improved bone formation and vascularisation activity. Markers associated with extracellular matrix (ECM) remodelling, including MMP9 and TIMP1, were also upregulated, suggesting active tissue remodelling. ELISA analysis for VEGF further demonstrated increased VEGF secretion, highlighting the scaffold’s angiogenic potential. The improved cellular response and vascular signalling emphasise the translational relevance of laser-modified GreenBone scaffolds for bone tissue engineering, particularly for critical-sized defect repair requiring rapid vascularised bone regeneration. Full article

Corilagin is a hydrolyzable ellagitannin and naturally occurring polyphenolic compound widely distributed in medicinal plants. It is also present in longan (Dimocarpus longan), known as lumyai in Thailand, a subtropical fruit extensively cultivated across China and Southeast Asia. Corilagin has been reported to exhibit strong antioxidant, anti-inflammatory, hepatoprotective, and anticancer activities through modulation of multiple cellular signaling pathways. However, despite these well-established pharmacological properties, its potential role in regulating bone marrow mesenchymal stem cell (BM-MSC) differentiation has not been fully explored in biomedical applications. In this study, we investigated the effects of corilagin on BM-MSC viability, protein-binding interactions, and lineage-specific differentiation toward osteogenic and chondrogenic pathways. Cytotoxicity assessment using human synovial SW-982 cells demonstrated that corilagin maintained cell viability at concentrations ranging from 1.56 to 50 µg/mL within 48 h, whereas prolonged exposure resulted in a time-dependent reduction in viability. In BM-MSCs, corilagin significantly enhanced osteogenic and chondrogenic differentiation in a dose-dependent manner, as evidenced by increased mineral deposition and cartilage matrix formation, as revealed by Alizarin Red S, Toluidine Blue, and Alcian Blue staining. Quantitative analyses further showed the upregulation of key lineage-specific genes, including Runx2 and osteopontin (OPN) for osteogenesis and Sox9 and aggrecan for chondrogenesis. Protein-binding assays confirmed the molecular interaction capacity of corilagin, supporting its biological activity. Overall, these findings demonstrate that corilagin promotes MSC-mediated osteogenic and chondrogenic differentiation while maintaining acceptable cytocompatibility, highlighting its potential as a natural small-molecule candidate for bone and cartilage tissue engineering and other biomedical fields with regenerative medicine applications. Full article

Kynurenic acid (KYNA), a small molecule derived from the tryptophan–kynurenine pathway, can readily diffuse across biological membranes and act as an endogenous ligand for receptors such as the aryl hydrocarbon receptor (Ahr). While KYNA dysregulation is implicated in neurodegenerative disorders, the role of the KYNA–Ahr-IL-6 axis in MSC proliferation and differentiation remains poorly defined. We investigated the impact of KYNA on murine bone marrow-derived MSCs (BM-MSCs) at various concentrations (10–200 μM) and time points (8–48 h). The BM-MSC phenotype was assessed via flow cytometry; proliferation, via cell counting; and the gene expression of Ahr, Cyp1a1, Cyp1b1, and Il-6, via quantitative real-time PCR. Multipotency was evaluated through adipogenic, osteogenic, and chondrogenic differentiation assays with histochemical confirmation. KYNA significantly upregulated Ahr mRNA expression. Among the tested concentrations, 100 μM KYNA induced the highest Ahr expression (~19.1 ± 1.5-fold greater than that of the untreated controls, p < 0.005). Notably, 10 and 50 μM KYNA caused moderate induction, whereas compared with 100 μM KYNA, 200 μM did not further increase expression. In addition, KYN treatment increased Cyp1a1, Cyp1b1, and Il-6 expression, with increases of ~64.6 ± 4.5-fold, ~43.6 ± 2.3-fold, and ~41.6 ± 1.2-fold, respectively. Compared with no treatment, 100 µM KYNA enhanced BM-MSC proliferation by 1.210 ± 0.02, 1.189 ± 0.03, and 1.242 ± 0.02-fold across passages P3, P4, and P5, respectively (p < 0.05), without altering Sca-1, CD90, or CD45 expression or impairing trilineage differentiation potential. KYNA may activate the AHR–IL-6 signaling axis to promote BM-MSC expansion. This controlled proliferative effect, without loss of phenotypic or functional integrity, highlights the pharmacological potential of KYNA as a small-molecule modulator for stem cell-based therapies. Full article

Mesenchymal stromal cells (MSCs) are multipotent and play an important role in regenerative processes such as wound healing. Data on possible changes and functional restrictions of MSCs due to cisplatin chemotherapy, for example, in the treatment of head and neck cancer, diverge. The aim of this study was to evaluate the influence of cisplatin on MSCs with regard to their defining characteristics and their ability to differentiate and to migrate. MSCs from four human donors (a 59-year-old man, a 63-year-old woman, a 70-year-old man, and a 61-year-old man) were cultured in vitro with and without cisplatin for 24 h, and toxic and subcytotoxic concentrations were determined using an MTT. We then examined the surface phenotype markers (flow cytometry), migration (scratch assay), histological differentiation markers (adipo-, chondro-, osteogenic), and the expression of selected line-associated genes in real-time quantitative PCR (RT-qPCR) (LEP, SOX9, RUNX2). These characteristics were evaluated after treatment with different subcytotoxic, clinically relevant doses of cisplatin. Flow cytometry confirmed the presence of MSCs-characteristic surface markers, which remained stable under treatment with subcytotoxic doses of cisplatin. Cisplatin exposure reduced the mRNA abundance of leptin (a marker for adipogenic differentiation) but increased SOX9 mRNA abundance (chondrogenic differentiation). RUNX (osteogenic differentiation) did not change post cisplatin exposure. Histological analysis showed no difference with regard to osteogenic, chondrogenic, and adipogenic differentiation at doses up to 10 μM cisplatin. Cell migration was not restricted by cisplatin exposure under the conditions used here. The characteristics of MSCs were not different to controls post cisplatin exposure. mRNA analysis suggested induced changes by cisplatin, although this effect was not histologically detectable even at high doses. Based on the single-molecule markers used here, indications for an inhibitory effect of cisplatin on adipogenic differentiation and a rather enhancing effect on chondrogenic and osteogenic differentiation may be hypothesized. The process observed here could further aggravate the already serious problem of malnutrition in head and neck cancer patients, for example. Taken together though, our study confirms overall MSCs tolerance towards cisplatin. Full article

Mesenchymal stem cells derived from human exfoliated deciduous teeth (SHED) are a promising source for regenerative therapies due to their multipotency, proliferative capacity, and immunomodulatory properties. The present study aimed to isolate and characterize SHED subpopulations based on CD71 and CD146 expression and evaluate their multilineage differentiation potential. SHED were obtained from pediatric donors and separated into CD71⁺, CD71⁻, CD146⁺, and CD146⁻ fractions using magnetic-activated cell sorting (MACS). CD71⁺/CD71⁻ and CD146⁺/CD146⁻ populations were isolated independently; no simultaneous double sorting for both markers was performed. Immunocytochemistry was employed to confirm the expression of surface and intracellular markers, including STRO-1, CD44, nestin, and vimentin. Multilineage differentiation assays toward osteogenic, adipogenic, and chondrogenic lineages revealed that CD71⁺ cells exhibited reduced osteogenic capacity compared to CD71⁻ cells, whereas CD146⁺ cells showed enhanced osteogenic and adipogenic differentiation. Chondrogenic differentiation seemed unaffected by marker expression under the 2D conditions employed. These results highlight functional heterogeneity within SHED populations and indicate that CD71 and CD146 independently influence differentiation outcomes. The selective enrichment of CD146⁺ SHED may enhance osteogenic and adipogenic regenerative applications, while CD71⁺ subsets may serve as a valuable model for studying proliferation and paracrine effects. Limitations include the use of in vitro differentiation assays and the absence of in vivo validation; additionally, combined CD71/CD146 analysis may further clarify the relationship between metabolic activity and stem/progenitor niche characteristics. Overall, marker-based characterization of SHED subpopulations provides insight into their biological properties and potential utility in targeted cell-based therapeutic strategies. Full article

(1) Objective: This study aimed to investigate the synergistic effect and underlying mechanisms of Total Flavonoids of Rhizoma drynariae (TFRD) in combination with Bone Marrow Mesenchymal Stem Cells (BMSCs) in the repair of tendon–bone injuries. (2) Methods: The effects of TFRD on the proliferation and migration of BMSCs were assessed using CCK-8 and scratch assays, and its potential to promote osteogenic and chondrogenic differentiation was evaluated. Concurrently, the pro-angiogenic effect of TFRD on Human Umbilical Vein Endothelial Cells (HUVECs) was observed. In vivo, a rat model of Achilles tendon–bone injury was established and animals were divided into four groups: SHAM, Model, BMSCs, and BMSCs + TFRD. After an 8-week intervention, the level of functional recovery was evaluated through histological analysis, immunohistochemistry, serum biochemical analysis, and biomechanical testing. (3) Results: A concentration of 5.0 μg/mL TFRD significantly promoted the proliferation, migration, and differentiation of BMSCs and enhanced the tube formation capacity of HUVECs. In the BMSCs + TFRD group, histological analysis revealed well-organized collagen fibers, increased cartilage deposition, and an optimized tendon–bone interface (TBI) structure. Immunohistochemistry showed upregulated expression of COL I, COL II, and SOX-9, alongside downregulated VEGFA. Furthermore, serum IL-6 levels were decreased, while IL-10 and TGF-β levels were elevated. The biomechanical properties were also significantly improved in this group. (4) Conclusions: TFRD promotes tendon–bone healing and functional recovery by enhancing BMSC functions, promoting angiogenesis, and improving the local microenvironment. Full article

Over the past decade, interest has grown in understanding the morphofunctional changes that mesenchymal stem cells (MSCs) undergo due to age-associated senescence—a process particularly relevant given that adults and elderly individuals are the primary candidates for regenerative therapies. This study addresses this knowledge gap by systematically analyzing the influence of age-related senescence on the morphofunctional properties of MSCs derived from the oral cavity. A scoping review was conducted following the PRISMA-ScR guidelines. The databases searched were MEDLINE, SCOPUS, and Web of Science. In vitro studies were included if their primary objective was to investigate oral cavity mesenchymal stromal cells and age-related senescence. A total of 455 studies were identified, of which 17 were selected. Studies on MSCs from the oral cavity have shown that age-related senescence, starting around 35 years, reduces proliferation, viability, clonogenic capacity, and differentiation potential—particularly toward osteogenic and chondrogenic lineages—with higher values observed in younger individuals. However, MSC surface markers remain stably expressed and show no association with aging. Some studies also report no significant differences in proliferation rate or cell doubling time at early passages, and MSCs retain some plasticity at these stages. Despite age-related limitations, oral MSCs from elderly donors remain a promising therapeutic source, especially at early in vitro passages. Further research is needed to explore innovative strategies to enhance the regenerative potential of oral MSCs from older donors. Full article

The objective of this review was to critically evaluate the available literature on the development of periodontal ligament organoids. Articles concerning periodontal ligament organoids were considered eligible. References were selected in a two-phased process. Electronic databases PubMed and Scopus were screened up to June 2024, yielding 1101 studies. After removing duplicates, titles, and abstracts were screened, resulting in 44 articles being included in this review. A detailed analysis of the included articles was organized into four categories: (1) the cell lineages used, including the simultaneous use of two or more cell types, (2) the extracellular matrix composition, (3) the organoid preparation methods, and (4) the characterization techniques employed. The main findings show that collagen combined with biodegradable polymers—such as poly(caprolactone), poly(glycolic acid), and poly(lactic acid)—is the most used material. Periodontal ligament cells and periodontal fibroblasts were the most used cell types, due to their role in extracellular matrix remodeling. The most frequent analyses performed included alkaline phosphatase, extracellular matrix mineralization, and gene expression, providing insights into differentiation and periodontal regeneration. Cementogenic differentiation was the most studied, followed by osteogenic, chondrogenic, adipogenic, and epithelial differentiation. However, challenges remain, including methodological inconsistencies and the need for scaffold optimization. Future research should focus on standardizing protocols, improving biomaterials, and integrating bioprinting techniques to improve clinical translation. Full article

Primary cells better reflect the physiological situation, and mesenchymal stromal cells (MSCs), especially, are promising candidates for biomedical applications. MSCs from the umbilical cord (UC) can be collected easily, non-invasively, and painlessly and do not involve ethical problems. The derived cell products harbor great potential in stem cell technology and agricultural applications. A tissue grinder (TIGR) was used to homogenize porcine UC tissue and to dissociate the UC cells, thereby testing different tissue-to-medium ratios. Cells were cultivated until passage 3, and the proliferation rate, metabolic activity, colony forming ability, surface marker expression, and multi-lineage differentiation potential were assessed. Tissue grinding could be successfully used to isolate UC-derived porcine cells with a high yield and viability, as well as an increasing proliferation rate during cultivation. Isolated cells showed MSC-like features: the expression of CD73, CD90, and CD105, ability to form colonies, and adipogenic, chondrogenic, and osteogenic differentiation. Tissue grinding is highly suitable for isolating high-quality cells from whole UC tissue of pigs in a fast and reproducible way. Cells might be used in a wide range of therapeutical and biotechnological applications, such as understanding and treating severe disorders, drug screening, or tissue engineering. Cells from supposedly waste tissues like UC will be especially useful in transplantation medicine. Full article