Search Results (362)

Background/Objectives: Chronic wounds represent a significant clinical and public health challenge due to impaired tissue repair and high associated morbidity. This study investigates the therapeutic potential of the secretome derived from human mesenchymal stem cells obtained from the pulp of deciduous teeth (hDP-MSCs) in promoting skin wound healing. Methods: After confirming the mesenchymal identity and multipotent differentiation potential of hDP-MSCs by using flow cytometry and histological staining, the effects of the secretome on human keratinocyte (HaCaT) cultures were evaluated. Results: Scratch assays, performed under high- and low-glucose conditions, demonstrated that the secretome significantly promoted keratinocyte migration and wound closure without compromising cell viability. Additionally, the secretome modulated the expression of key genes involved in inflammation and tissue regeneration, including IL-1β, TNF-α, TGF-β1, and VEGF-α, in a time-dependent manner. Under inflammatory conditions induced by lipopolysaccharide, co-treatment with the secretome significantly reduced TNF-α expression and increased TGF-β1 expression, suggesting an anti-inflammatory effect. Conclusions: These findings indicate the potential of the hDP-MSC-derived secretome as a promising cell-free therapeutic strategy capable of accelerating skin regeneration and modulating the inflammatory response during the wound healing process. Full article

Cutaneous wound healing is a complex process involving multiple cellular and molecular events, and current treatments often face limitations in efficacy and safety. Stem-cell therapy, particularly using mesenchymal stem cells (MSCs), has emerged as a promising approach to enhance wound repair through both direct cell replacement and paracrine signaling. This study investigates the therapeutic potential of human chorionic villus mesenchymal stem cells (hCV-MSCs) and their secretory factors in enhancing cutaneous wound healing. Utilizing a rat model, we combined the local administration of hCV-MSC-laden PEGDA/SA/Col-I hydrogel with the systemic delivery of their secretome, aiming to leverage the complementary mechanisms of cellular and cell-free therapies. Our findings demonstrate that hCV-MSCs delivered via PEGDA/SA/Col-I hydrogel significantly accelerated wound closure compared to controls, with near-complete closure observed by day 20. Histological analysis revealed enhanced keratinocyte maturation (increased KRT10/KRT14 ratio) and a higher density of CD31⁺ blood vessels, indicating improved re-epithelialization and angiogenesis. A mass spectrometry analysis of the hCV-MSC secretome identified 849 proteins, with enrichment in pathways related to ECM organization, cell adhesion, and immune regulation. Key proteins such as ANXA1, SERPINE1, and WNT5A were implicated in wound-healing processes. Combination therapy with systemic secretome administration further accelerated wound closure and enhanced collagen deposition, keratinocyte maturation, and vascularization compared to hCV-MSCs alone. Our results highlight the promising application of hCV-MSCs and their secretome in cutaneous wound healing, paving the way for innovative therapeutic strategies that integrate both local and systemic regenerative approaches. Full article

Nerve tissue damage is an unsolved problem in modern neurology and neurosurgery, which prompts the need to search for approaches to stimulate neuroprotection and regeneration of neural tissue. Earlier we have shown that the secretome of human mesenchymal stromal cells (MSCs) stimulates rat survival, reduces the severity of neurological deficits, and decreases the volume of brain damage in a hemorrhagic stroke model. A significant disadvantage of using the MSC secretome is the need to concentrate it (at least 5–10 fold) to achieve appreciable pharmacological activity. This increases the cost of obtaining clinically applicable amounts of secretome and slows down the clinical translation of this technology. Here, we created a number of genetically modified human MSC cultures, including immortalized MSCs and those with hyperexpression of brain-derived neurotrophic factor (BDNF) and urokinase-type plasminogen activator (uPA) and with suppressed expression of Von Hippel–Lindau tumor suppressor (VHL), and we evaluated the pharmacological activity of their secretomes in a model of intracerebral hemorrhage (ICH) in rats. The secretome of MSCs immortalized by hyperexpression of the catalytic subunit of human telomerase (hTERT) revealed neuroprotective activity indistinguishable from that of primary MSC cultures, yet it still required 10-fold concentration to achieve neuroprotective efficacy. The secretome of MSC culture with combined hyperexpression of BDNF and uPA and suppressed expression of Von Hippel–Lindau tumor suppressor even without additional concentration reduced the severity of neurological disorders and decreased brain lesion volume in the ICH model. The secretomes of MSCs with separate overexpression of BDNF and uPA or suppression of VHL had no such effect or, on the contrary, revealed a toxic effect in the ICH model. Presumably, this may be due to an imbalance in the representation of individual growth factors in the secretome of genetically modified MSCs, which individually may lead to undesirable effects in damaged nervous tissue, such as increased permeability of the blood–brain barrier (under the influence of pro-angiogenic factors) or neural cell apoptosis (due to an excess of neurotrophic factors). The obtained data show that genetic modification of MSC cultures can enhance or alter the therapeutic activity of their secretomes, which can be used in the creation of promising sources of biopharmaceutical substances. Full article

Mesenchymal stem cells (MSCs) represent a promising cell source for cartilage tissue engineering due to their chondrogenic potential. However, current differentiation protocols result in limited efficiency. This study assessed the combined effects of transforming growth factor-beta 3 (TGF-β3) and fibroblast growth factor-2 (FGF-2) on the morphology, proliferation, chondrogenic differentiation, chondrogenic gene expression, and cytokine profile of ovine bone marrow-derived MSCs (BM-MSCs). BM-MSCs were cultured under four conditions: control (αMEM) or αMEM supplemented with FGF-2, TGF-β3, or TGF-β3 + FGF-2. Morphological and proliferation analyses, Alcian blue staining in 2D and 3D, and real-time PCR for early (Chad, Comp, and Sox 5) and late (Agg, Col IX, Sox 9, and Fmod) chondrogenic markers were performed. Cytokine secretion profiles were analyzed using multiplex assay. TGF-β3 induced morphological changes indicative of early chondrogenesis, while FGF-2 enhanced proliferation. The combination of both cytokines led to a synergistic increase in cell proliferation, early and late chondrogenic gene expression, and glycosaminoglycans (GAG) deposition. Cytokine analysis revealed that TGF-β3 enhanced the immunomodulatory and angiogenic profile of BM-MSCs, whereas co-treatment with FGF-2 yielded a balanced and potentially regenerative secretome. Dual stimulation with TGF-β3 and FGF-2 significantly improves the chondrogenic differentiation of ovine BM-MSCs by enhancing both molecular and functional markers of cartilage formation. Full article

Skin aging is mostly caused by the accumulation of reactive oxygen species (ROS) that lead to cellular dysfunction. One promising therapy for skin aging is the secretome product of adipose tissue mesenchymal stem cells (AT-MSCs). L-ascorbic acid (LAA) is an essential molecule for preventing oxidative stress as an external antioxidant agent and has been used in chemical therapy for skin aging. In this study, we evaluated the effects of LAA on cell morphology, the number of cells, cell viability, and the paracrine secretion of preconditioned AT-MSCs in in vitro culture with LAA in 100 and 200 µg/mL compared with an untreated culture with LAA as a control. LAA supplementation in both concentrations improved the morphology of cells without affecting the cell viability. However, there was no significant improvement in the number of cells even though the trend showed an enhancement of the number of cells. The total protein of the secretome decreased in the LAA preconditioning group. However, preconditioning AT-MSCs in in vitro culture with LAA improved the levels of insulin-like growth factor 1 (IGF-1), transforming growth factor β1 (TGF-β1), and interleukin 6 (IL-6) which are essential proteins for skin aging in regulating ROS. Full article

Background/Objectives: Mesenchymal stem cells (MSCs) possess immunomodulatory properties, tumor-homing, and low immunogenicity, making them attractive for cell-based cancer therapies, but their role in colorectal cancer (CRC) remains controversial. The MSC1 phenotype, a pro-inflammatory, tumor-suppressive state induced by short-term, low-dose LPS activation via TLR4, has shown therapeutic promise but remains poorly characterized in CRC. We aimed to elucidate MSC1’s tumor-suppressive mechanisms and validate its activity against CRC cells using an integrated bioinformatics and in vitro approach. Methods: We constructed a high-confidence protein-protein interaction (PPI) network in Wharton’s jelly-derived MSCs (WJ-MSCs) following TLR4 activation to uncover enriched signaling pathways, transcriptional regulators, and secreted factors. Functional and transcriptional enrichment analyses pinpointed key mechanisms. We then co-cultured MSC1 cells with CRC cells to assess effects on proliferation and metabolism. Results: Network analysis revealed six tumor-suppressive mechanisms of MSC1 cells: (i) Metabolic reprogramming via enhanced glucose and lipid uptake, phosphoinositide signaling, and membrane/protein recycling, (ii) Robust antioxidant defenses, including SOS signaling and system xc⁻, (iii) Extracellular matrix stabilization and laminin-111–integrin-mediated adhesion, (iv) Secretome with direct anti-cancer effects, (v) Regulation of survival and cancer-associated fibroblasts (CAFs) formation inhibition through balanced proliferation, apoptosis, and epigenetic signals, (vi) Controlled pro-inflammatory signaling with anti-inflammatory feedback. In vitro, MSC1 cells significantly suppressed CRC cell proliferation and metabolic activity versus controls. Conclusions: This study provides the first mechanistic map of MSC1’s tumor-suppressive functions in CRC, extending beyond immunomodulation to include metabolic competition, ECM stabilization, and anti-cancer secretome activity. These findings establish MSC1 cells as a novel therapeutic strategy for CRC in cell-based cancer therapies. Full article

Systemic cancer progression culminating in metastatic disease is implicitly dependent on tumour cell interactions with the vascular system. Indeed, different facets of the micro- and macro-vasculature can be regarded as rate-limiting ‘vascular checkpoints’ in the process of cancer dissemination. The underlying complex communication networks drive tumour neovascularization, angiogenesis, immunoregulation, activation of the coagulation system, angiocrine interactions, and non-angiogenic vascular responses across multiple cancer types. Yet, each cancer may represent a unique vascular interaction scenario raising a prospect of targeted modulation of blood and lymphatic vessels for therapeutic purposes, beyond the traditional notion of tumour anti-angiogenesis. While the emphasis of studies aiming to understand this circuitry has traditionally been on soluble, or ‘mono-molecular’ mediators, the rise of the particulate secretome encompassing heterogeneous subpopulations of extracellular vesicles (EVs; including exosomes) and particles (EPs) brings another dimension into the tumour–vascular communication web during the process of metastasis. EVs and EPs are nanosized cellular fragments, the unique nature of which lies in their ability to encapsulate, protect and deliver to target cells a range of bioactive molecular entities (proteins, RNA, DNA) assembled in ways that enable them to exert a wide spectrum of biological activities. EVs and EPs penetrate through biological barriers and are capable of intracellular uptake. Their emerging vascular functions in metastatic or infiltrative cancers are exemplified by their roles in pre-metastatic niche formation, thrombosis, vasectasia or angiocrine regulation of cancer stem cells. Here, we survey some of the related evidence supporting the biological, diagnostic and interventional significance of EVs/EPs (EVPs) in disseminated neoplastic disease. Full article

Tendon ruptures have recently reached incidences of 18–35 cases/100,000 and often lead to adhesion formation during healing. Furthermore, scar formation may result in inferior biomechanics and often leads to re-ruptures. To address these problems, we cultivated rabbit adipose-derived stem cells in a co-culture with rabbit Achilles tenocytes and harvested their secretome. Following a cell-free approach, we incorporated such secretome into an electrospun tube via emulsion electrospinning. These novel implants were characterized by SEM, the WCA, and FTIR. Then, they were implanted in the rabbit Achilles tendon full transection model with an additional injection of secretome, and the adhesion extent as well as the biomechanics of extracted tendons were assessed three weeks postoperatively. The fiber thickness was around 3–5 μm, the pore size 11–13 μm, and the tube wall thickness approximately 265 μm. The WCA indicated slightly hydrophilic surfaces in the secretome-containing layer, with values of 80–90°. In vivo experiments revealed a significant reduction in adhesion formation (−22%) when secretome-treated tendons were compared to DegraPol^® (DP) tube-treated tendons (no secretome). Furthermore, the cross-sectional area was significantly smaller in secretome-treated tendons compared to DP tube-treated ones (−32%). The peak load and stiffness of secretome-treated tendons were not significantly different from native tendons, while tendons treated with pure DP tubes exhibited significantly lower values. We concluded that secretome treatment supports tendon healing, with anti-adhesion effects and improved biomechanics at 3 weeks, making this approach interesting for clinical application. Full article

Prion diseases are disorders caused by the misfolding of prion protein (PrP^Sc), leading to the accumulation of an abnormal form of the normal prion protein (PrP) found in the host. The secretome of mesenchymal stem cells (MSCs), including paracrine-soluble factors, holds promising potential to stimulate host regenerative capability and alleviate organ disorders. In this research, our goal was to investigate the neuroprotective properties of the secretome derived from adipose-derived mesenchymal stem cells (AdMSC secretome) in relation to the toxicity caused by PrP^106−126 in SH-SY5Y cells. The findings showed that PrP^106−126 treatment exacerbated the neurotoxicity of SH-SY5Y cells, as indicated by increased lactate dehydrogenase (LDH) release. However, the AdMSC secretome significantly decreased LDH release. Under PrP^106−126 stimulation, the AdMSC secretome downregulated inflammatory markers (TNF-α and IL-1β) and upregulated anti-inflammatory IL-10. Treatment with the AdMSC secretome markedly reduced GFAP immunoreactivity in astrocytic C8D1A cells compared to treatment with PrP^106−126 alone. In addition, the AdMSC secretome reduced Iba-1 immunoreactivity in BV2 cells activated by LPS. Western blot analysis showed that the AdMSC secretome inhibited pro-apoptotic factor Bax induced by PrP^106−126 and increased the expression of anti-apoptotic factor Bcl-2. However, no significant difference was observed in the expression of caspase-3. The AdMSC secretome exhibited a considerable migratory effect on SH-SY5Y cells after 24 h, as demonstrated by the scratch assay. The results suggest that the AdMSC secretome can attenuate PrP^106−126-induced neuronal damage. Full article

Stem cell-derived secretome and exosomes present a promising cell-free strategy for tissue repair and wound healing. This study aimed to isolate and characterize, for the first time, exosomes derived from rat hair follicle stem cells (rHFSCs) and to evaluate their wound-healing potential alongside rHFSC secretome. Exosomes were isolated via ultracentrifugation and characterized using Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), biomarker profiling and protein quantification. Scanning electron microscopy (SEM) with energy-dispersive X-ray spectroscopy (EDS) confirmed their spherical morphology, diameter and elemental composition. Protein quantification showed higher protein content in the secretome than in exosomes. RT-PCR and biomarker profiling highlighted the therapeutic relevance of the exosomal cargo compared to parent rHFSCs. Functional analysis of 30 wound-healing biomolecules validated their pro-regenerative potential. Cytocompatibility was confirmed via the PrestoBlue™ viability assay, while scratch assays demonstrated significant wound closure in the treated groups, both with and without mitomycin C. These findings highlight the potential of rHFSC-derived exosomes and secretome as innovative, cell-free therapeutic agents for cutaneous regeneration. This study advances our understanding of their role in wound healing and underscores their broader applicability in regenerative medicine. Full article

Parkinson’s disease (PD) is a neurodegenerative disorder primarily defined by the deterioration of motor function and characterized by the loss of dopaminergic neurons in the nigrostriatal system. Although it is the second most prevalent disorder of the central nervous system, current treatments primarily focus on symptom management and modestly slowing disease progression, ultimately failing to preserve the long-term quality of life of a substantial proportion of affected individuals. Innovative therapies that can restore neuronal function have emerged, such as the use of the secretome of Mesenchymal Stem Cells (MSCs) due to their rich composition of bioactive molecules. This therapy exhibits robust paracrine activity that drives most of the self-renewal capacity, differentiation potential, and immune regulation of MSCs without presenting compatibility issues often associated with stem cell-based therapies. While conceptually appealing, the clinical application of this approach is still limited by the availability and proliferation capacity of MSCs, as it impacts not only secretome production but also its quality. Various protocols have been developed to enhance secretome action by adding various compounds to cell culture media, given the high environmental plasticity of MSCs. Some of the compounds already used are Caloric Restriction Mimetics (CRMs), molecules that mimic Caloric Restriction (CR) conditions, which have been demonstrated to extend lifespan and reduce age-related diseases in various organisms. While not sufficient to cure neurodegenerative disorders, these compounds may potentiate secretome efficiency by enhancing autophagy pathways and relieving oxidative stress burden from MSCs. Therefore, in this article, we aim to explore the effects of CRMs priming on MSCs and how it may help bridge existing gaps in regenerative therapies for PD. Full article

Treating endometritis is one of the major challenges faced by veterinarians. The effect of post-insemination intrauterine infusion Wharton’s jelly mesenchymal stromal/stem cell-derived conditioned medium (WJ-MSC-CM) on the uterine response and pregnancy outcomes in problem mares was evaluated in this study. The cycles of 12 problem mares were treated seven/eight hours after insemination with intrauterine infusion of 20 mL of WJ-MSC-CM (TRT) or non-conditioned Ringer’s lactate solution (CTR). Before (PRE) and 12 h after (POST) treatment, an ultrasonographic examination and a low-volume flush (LVF) for efflux clarity evaluation and bacteriological and cytological analysis were made. Pregnancy diagnosis was performed at 14 days and confirmed at 60 days after ovulation. A decrease (p < 0.05) in polymorphonuclear neutrophils infiltration and a trend toward significance in LVF efflux clarity (p < 0.1) were observed in TRT cycles. The TRT (50%) had a higher pregnancy rate (p < 0.05) than CTR group (0%) at first cycle. Only a trend toward significance was observed comparing pregnancy frequencies between the two groups (CTR 12.5% vs. TRT 54.5%), the pregnancy rate of TRT cycles was comparable to that of normal mares (p > 0.05) and higher (p < 0.05) than that reported for problem mares in the literature. Post-insemination uterine infusion of WJ-MSC-CM in mares restores pregnancy rates to normal levels, by its anti-inflammatory effect. Full article

Background: Mesenchymal stem cell-based therapy may be indicated in ischaemic heart disease. The use of autologous adipose-derived mesenchymal stromal cells (AdMSCs) offers regenerative potential due to their paracrine effects. The aim of this study was to expand and characterise adult human thymus-derived MSCs harvested during open heart surgery with respect to their stem cell and paracrine properties. Methods: Enzymatically and non-enzymatically isolated human thymic AdMSCs (ThyAdMSCs) were cultured in xeno-free media containing pooled human platelet lysate (pPL). MSC characterisation was performed. Ex vivo expanded ThyAdMSCs were differentiated into three lineages. Proliferative capacity and immunomodulatory properties were assessed by proliferation assays and mixed lymphocyte reaction, respectively. Gene expression analysis was performed by qPCR. Results: Both isolation methods yielded fibroblast-like cells with plastic adherence and high proliferation. Flow cytometry revealed distinct expression of MSC markers in the absence of haematopoietic cell surface markers. Ex vivo expanded ThyAdMSCs could be differentiated into adipocytes, osteocytes, and chondrocytes. Activated peripheral blood mononuclear cells were significantly reduced when co-cultured with ThyAdMSCs, indicating their ability to inhibit immune cells in vitro. Gene expression analysis showed significantly less IFNγ and TNFα, indicating an alteration of the activated and pro-inflammatory state in the presence of ThyAdMSCs. Conclusions: These results demonstrate an efficient method to generate AdMSCs from human thymus. These MSCs have a strong immunomodulatory capacity and are, therefore, a promising cell source for regenerative medicine. The culture conditions are crucial for cells to proliferate in culture. Further research could explore the use of ThyAdMSCs or their secretome in surgical procedures. Full article

Extensive research has been conducted on mesenchymal stem cells (MSCs) regarding their ability to modify the immune response and reduce tissue damage. Many researchers have found that the regulatory capacity of MSCs primarily comes from their secretome. As a result, there has been much interest in utilizing “cell-free” therapies as alternatives to stem cell treatments. In this study, the secretome from adipose mesenchymal stem cells (ADSC-secretome) was extracted and injected into minipigs with established liver injury models. Blood and liver tissue samples were obtained prior to the procedure, as well as on days 1, 3, and 7 after surgery. It was found that ADSC-secretome effectively suppressed the synthesis of the NOD-like receptor protein 3 (NLRP3) inflammasome, leading to a downregulation of gasdermin-D (GSDMD) expression, and demonstrated a more prominent anti-pyroptosis effect compared to ADSCs. Furthermore, ADSC-secretome inhibited the high mobility group box 1 (HMGB1)/toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) inflammatory pathway. In summary, both ADSC-secretome and ADSCs inhibited pyroptosis in right hemihepatic ischemia–reperfusion combined with left hemihepatectomy injury, and ADSC-secretome exhibited a stronger therapeutic effect. ADSC-secretome exerted these therapeutic effects through the inhibition of the HMGB1/TLR4/NF-κB inflammatory pathway. In the future, “cell-free” therapy is expected to replace cell-based methods. Full article

Objective: Secretory factors, termed the secretome, in the conditioned medium (CM) from dental mesenchymal stem cells (MSCs) have shown anti-inflammatory, anti-apoptotic, and tissue regenerative potential. This cell-free product could be further developed by preconditioning cells with various biochemical agents, which lead to a change in secretome and CM profiles. Among the favorable candidates for CM production, metformin as an anti-diabetic medication is currently considered a potential agent for dental hard tissue and periodontal regeneration. Here, we aimed to assess the composition of CM from periodontal ligament stem cells (PDLSCs) grown in metformin-preconditioned media (Met-CM) compared to normal PDLSC-CM and assess the ability of Met-CM to recover the function of inflamed PDLSCs. Methods: Met-CM and normal CM were collected from PDLSCs grown with or without 50 µM metformin, respectively, under healthy culture conditions. Mass spectrometry and liquid chromatography–tandem mass spectrometry (LC–MS/MS) were performed to comparatively evaluate the proteomic profiles in PDLSC-CM versus Met-CM. We then treated the PDLSC cultures with lipopolysaccharide (LPS) from Porphyromonas gingivalis to induce inflammation and evaluated the osteogenic/cementogenic differentiation in the presence of Met-CM or normal PDLSC-CM by assessing alkaline phosphatase activity, intracellular calcium levels, and mRNA expression of osteogenic and cementogenic factors, including RUNX2, OCN, OSX, and CEMP-1. Subsequently, we performed RNA sequencing to identify transcriptomic changes in the treated cells. Results: We identified 202 differentially expressed proteins, 175 of which were significant, in Met-CM versus normal PDLSC-CM. Among the analyzed groups, the top three protein classes were protein-binding activity modulator, cytoskeletal protein, and extracellular matrix (ECM) protein. Treatment of PDLSCs with LPS significantly attenuated ALP activity, [Ca²⁺]_i, and the mRNA expression levels of RUNX2, OCN, OSX, and CEMP-1, whereas treatment with Met-CM alone markedly enhanced PDLSC differentiation activity compared with the control. Moreover, osteogenic/cementogenic differentiation of the LPS-treated PDLSCs was recovered through incubation in Met-CM. Transcriptomic analysis identified 511 and 3591 differentially expressed genes in the control versus Met-CM and LPS versus LPS + Met-CM groups, respectively. The enrichment of biological processes includes positive regulation of DNA-templated transcription and skeletal system morphogenesis in the control versus Met-CM comparison, as well as positive regulation of transcription from the RNA polymerase II promoter and negative regulation of the apoptotic process in the LPS versus LPS + Met-CM comparison. Molecular function analysis demonstrated the enrichment of protein-binding terms among the DEGs from each comparison. Conclusions: Metformin preconditioning enhanced the recovery effect of PDLSC-CM on LPS-induced inflamed PDLSCs. These findings suggest that metformin preconditioning could represent a practical formula for PDLSC-secretome, which may contribute to the development of future cell-free periodontal regenerative strategies. Full article