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Background aimsCulture-derived mesenchymal stromal cells (MSCs) exhibit variable characteristics when manufactured using different methods, source material and culture media. The purpose of this multicenter study was to assess the impact on MSC expansion, gene expression and other characteristics when different laboratories expanded MSCs from cultures initiated with bone marrow-MSC aliquots derived from the same donor source material yet with different growth media.MethodsEight centers expanded MSCs using four human platelet lysate (HPL) and one fetal bovine serum (FBS) products as media supplements. The expanded cells were taken through two passages then assessed for cell count, viability, doubling time, immunophenotype, cell function, immunosuppression and gene expression. Results were analyzed by growth media and by center.ResultsCenter methodologies varied by their local seeding density, feeding regimen, inoculation density, base media and other growth media features (antibiotics, glutamine, serum). Doubling times were more dependent on center than on media supplements. Two centers had appropriate immunophenotyping showing all MSC cultures were positive for CD105, CD73, CD90 and negative for CD34, CD45, CD14, HLA-DR. MSCs cultured in media supplemented with FBS compared with HPL featured greater T-cell inhibition potential. Gene expression analysis showed greater impact of the type of media supplement (HPL versus FBS) than the manufacturing center. Specifically, nine genes were decreased in expression and six increased when combining the four HPL-grown MSCs versus FBS (false discovery rate [FDR]

Background: A significant number of platelet concentrates (PCs) is discarded daily in blood banks due to limited shelf life. Human platelet lysate (HPL), derived from expired PCs, has gained attention as an ethical and sustainable cell culture media supplement in biomedical research and cell therapy production. However, HPL is subject to decisive disadvantages such as batch differences and lack of storage stability. To overcome these limitations and to enhance the applicability of HPL, we developed an HPL manufacturing protocol including a lyophilization process. The aim of this study was to investigate the influence of HPL lyophilization on parameters of quality control, including growth factor concentrations and the culture of human mesenchymal stromal cells (hMSCs). Methods: We performed a paired comparison of six batches of HPL and lyophilized HPL (L-HPL) regarding the quality parameters pH, total protein, osmolality, sodium, potassium and chloride concentration. Concentrations of 11 growth factors and cytokines were compared between HPL and L-HPL. Additionally, we determined cell yield, proliferation capacity, viability and trilineage differentiation potential of hMSCs following expansion in HPL- and L-HPL-supplemented cell culture media. Results: Quantification of the quality parameters revealed non-altered pH, osmolality and potassium concentrations and slightly lower total protein, sodium and chloride concentrations of L-HPL compared to HPL. Growth factor and cytokine concentrations did not differ between HPL and L-HPL. Cell yield, division cycles and viability of hMSCs cultured in either HPL- or L-HPL-containing media were comparable. Cells differentiated in medium containing L-HPL showed a slightly higher capacity for osteogenic differentiation, while adipogenic differentiation and chondrogenic differentiation potentials remained unchanged. Conclusion: We successfully developed a method to produce well-applicable L-HPL. The comparison of L-HPL with HPL did not reveal any relevant differences regarding quality control parameters of routine testing, growth factor concentrations and hMSC functionality, demonstrating the suitability of L-HPL as a cell culture supplement. These results emphasize the potential of L-HPL as a sustainable and ethical alternative to animal-derived serum products in biomedical research and drug development. [ABSTRACT FROM AUTHOR]

Background: Extracellular vesicles (EVs) secreted by mesenchymal stromal cells (MSCs) have been shown to provide significant protection against renal ischemia–reperfusion injury (IRI). Hypoxia has emerged as a promising strategy to enhance the tissue repair capabilities of MSCs. However, the specific effects of hypoxia on MSCs and MSC-EVs, as well as their therapeutic potential in renal IRI, remain unclear. In this study, we investigated the alterations occurring in MSCs and the production of MSC-EVs following hypoxia pre-treatment, and further explored the key intrinsic mechanisms underlying the therapeutic effects of hypoxic MSC-EVs in the treatment of renal IRI. Methods: Human umbilical cord MSCs were cultured under normoxic and hypoxic conditions. Proliferation and related pathways were measured, and RNA sequencing was used to detect changes in the transcriptional profile. MSC-EVs from both normoxic and hypoxic conditions were isolated and characterized. In vivo, the localization and therapeutic effects of MSC-EVs were assessed in a rat renal IRI model. Histological examinations were conducted to evaluate the structure, proliferation, and apoptosis of IRI kidney tissue respectively. Renal function was assessed by measuring serum creatinine and blood urea nitrogen levels. In vitro, the therapeutic potential of MSC-EVs were measured in renal tubular epithelial cells injured by antimycin A. Protein sequencing analysis of hypoxic MSC-EVs was performed, and the depletion of Glutathione S-Transferase Omega 1 (GSTO1) in hypoxic MSC-EVs was carried out to verify its key role in alleviating renal injury. Results: Hypoxia alters MSCs transcriptional profile, promotes their proliferation, and increases the production of EVs. Hypoxia-pretreated MSC-EVs demonstrated a superior ability to mitigate renal IRI, enhancing proliferation and reducing apoptosis of renal tubular epithelial cells both in vivo and in vitro. Protein profiling of the EVs revealed an accumulation of numerous anti-oxidative stress proteins, with GSTO1 being particularly prominent. Knockdown of GSTO1 significantly reduced the antioxidant and therapeutic effects on renal IRI of hypoxic MSC-EVs. Conclusions: Hypoxia significantly promotes the generation of MSC-EVs and enhances their therapeutic effects on renal IRI. The antioxidant stress effect induced by GSTO1 is identified as one of the most critical underlying mechanisms. Our findings highlight that hypoxia-pretreated MSC-EVs represent a novel and promising therapeutic strategy for renal IRI. [ABSTRACT FROM AUTHOR]

Introduction: Articular cartilage damage presents a significant clinical challenge, with limited options for effective regeneration. Mesenchymal stromal cells (MSCs) derived from Wharton's jelly (WJ) are a promising cell source for cartilage repair due to their regenerative and immunomodulatory properties. While undifferentiated MSCs have demonstrated potent immunoregulatory effects, the immunomodulatory potential of chondrocytes derived from WJ-MSCs remains underexplored, particularly under inflammatory conditions. This study investigates the differential cytokine expression profiles of WJ-MSC-derived chondrocytes and undifferentiated MSCs under inflammatory stimulation with phytohemagglutinin (PHA) to understand their immunomodulatory capacities. Materials and Methods: WJ-MSCs were differentiated into chondrocytes using a micromass culture system. Differentiated chondrocytes were then co-cultured with immune cells under PHA-induced inflammatory conditions. Control groups included co-cultured cells without PHA activation and chondrocytes activated with PHA in the absence of immune cell interaction. Cytokine expression profiles were analyzed using the RT2 Customized Gene Array to evaluate pro- and anti-inflammatory markers. Morphological changes were assessed microscopically. The immunomodulatory responses of chondrocytes were compared to those of undifferentiated MSCs under the same experimental conditions. Results: Chondrocytes co-cultured with immune cells under PHA activation exhibited downregulation of IDO, HLA-G, PDGF, IL-10, TNF-α, IL-6, and IFN-γ compared to undifferentiated MSCs in similar conditions. In non-PHA co-cultured conditions, chondrocytes showed increased expression of IL-6, IFN-γ, IL-4, VEGF, iNOS, PDGF, PTGS-2 and TGF-β, while TNF-α, IL-10, IDO and HLA-G were decreased. In contrast, chondrocytes activated with PHA without immune cell interaction displayed reduced expression of HLA-G and TNF-α, with no significant changes in IL-6, IFN-γ, IL-4, IL-10, VEGF, PDGF, PTGS-2, TGF-β, IDO, and iNOS compared to PHA-stimulated undifferentiated MSCs. Conclusion: This study demonstrates that chondrocytes derived from WJ-MSCs exhibit limited immunomodulatory potential compared to undifferentiated MSCs, particularly under PHA-induced inflammatory conditions. Undifferentiated MSCs showed superior regulation of key cytokines associated with immune modulation. These findings suggest that maintaining MSCs in an undifferentiated state may be advantageous for therapeutic applications targeting inflammatory conditions, such as osteoarthritis. Future research should explore strategies to enhance the immunomodulatory efficacy of chondrocytes, potentially through genetic modification or adjunctive therapies. [ABSTRACT FROM AUTHOR]

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is still a growing concern in the field of antimicrobial resistance due to its resistance to conventional antibiotics and its association with high mortality rates. Mesenchymal stromal cells (MSCs) have been shown as a promising and attractive alternative treatment for bacterial infections, due to their antibacterial properties and potential to bypass traditional resistance mechanisms. This study aims to shed light on the antibacterial potential of adipose-derived mesenchymal stromal cell (AD-MSC) secretome against clinical isolates of Staphylococcus spp., including MRSA strains. Methods: Using the Kirby-Bauer disk diffusion method, broth microdilution assays, and colony-forming unit (CFU) counting, the antibacterial activity of AD-MSC secretome was assessed. These tests were first conducted on Staphylococcus (S.) aureus ATCC 25923, then on 73 clinical isolates including MRSA strains. Further molecular analysis was performed to identify resistant genes in MRSA isolates. Results: The AD-MSC secretome demonstrated significant antibacterial activity against S. aureus ATCC with a 32 mm inhibition zone. 96% of the collected staphylococcal clinical isolates showed susceptibility to the secretome with 87.5% inhibition observed in MRSA isolates, along with 100% in MSSA, MSSE, and MRSE strains. Molecular analysis revealed that MRSA strains resistant to the secretome harbored mecA, ermA, and ermB genes. Additionally, the mecA-negative MRSA strains remained susceptible to the secretome, suggesting alternative resistance mechanisms. Conclusion: These findings emphasize the ability of AD-MSCs secretome as a promising alternative for treating antibiotic-resistant infections, with potential applications in combating MRSA. However, further research is required to explore its clinical applications as a complementary or standalone therapy for resistant infections. [ABSTRACT FROM AUTHOR]

Interest in prostate cancer as a research topic has gradually increased. As a result, a series of innovative treatment strategies have emerged with an in-depth understanding of the disease. Owing to their unique biological characteristics, mesenchymal stromal cell exosomes (MSC-Exos) have garnered significant attention for their potential to deliver targeted drugs and enable precise prostate cancer treatment. Herein, prostate cancer treatment with MSC-Exos drug-delivery systems is reviewed. This review provides a comprehensive introduction to the advantages of these systems, current research trends and progress, as well as an analysis of current challenges and future research directions. Moreover, this review lays a solid foundation for the continued development and application of MSC-Exos. [ABSTRACT FROM AUTHOR]

Objective: The inflammatory responses from synovial fibroblasts and macrophages and the mitochondrial dysfunction in chondrocytes lead to oxidative stress, disrupt extracellular matrix (ECM) homeostasis, and accelerate the deterioration process of articular cartilage in osteoarthritis (OA). In recent years, it has been proposed that mesenchymal stromal cells (MSC) transfer their functional mitochondria to damaged cells in response to cellular stress, becoming one of the mechanisms underpinning their therapeutic effects. Therefore, we hypothesize that a novel cell-free treatment for OA could involve direct mitochondria transplantation, restoring both cellular and mitochondrial homeostasis. Methods: Mitochondria were isolated from Umbilical Cord (UC)-MSC (Mito-MSC) and characterized based on their morphology, phenotype, functions, and their ability to be internalized by different articular cells. Furthermore, the transcriptional changes following mitochondrial uptake by chondrocytes were evaluated using an Affymetrix analysis, Lastly, the dose dependence therapeutic efficacy, biodistribution and immunogenicity of Mito-MSC were assessed in vivo, through an intra-articular injection in male C57BL6 mice in a collagenase-induced OA (CIOA) model. Results: Our findings demonstrate the functional integrity of Mito-MSC and their ability to be efficiently transferred into chondrocytes, synovial macrophages, and synovial fibroblasts. Moreover, the transcriptomic analysis showed the upregulation of genes involved in stress such as DNA reparative machinery and inflammatory antiviral responses. Finally, Mito-MSC transplantation yielded significant reductions in joint mineralization, a hallmark of OA progression, as well as improvements in OA-related histological signs, with the lower dose exhibiting better therapeutic efficacy. Furthermore, Mito-MSC was detected within the knee joint for up to 24 h post-injection without eliciting an inflammatory response in CIOA mice. Conclusion: Collectively, our results reveal that mitochondria derived from MSC are transferred to key articular cells and are retained in the joint without generating an inflammatory immune response mitigating articular cartilage degradation in OA, probably through a restorative effect triggered by the stress antiviral response within OA chondrocytes. [ABSTRACT FROM AUTHOR]

Background: As cell-free nanotherapeutics, extracellular vesicles derived from mesenchymal stem cells (MSC-EVs) have shown potential therapeutic action against liver diseases. However, their effects on autoimmune hepatitis (AIH) are not yet well understood. Methods and results: In this study, we utilized a well-established concanavalin A (Con A)-induced fulminant hepatitis mouse model to investigate the effects of MSC-EVs on AIH. We found that MSC-EVs provide significant protection against Con A-induced hepatitis in C57BL/6 male mice, with their effectiveness being critically dependent on the gut microbiota. MSC-EVs modulate the composition of the gut microbiota, particularly by increasing the abundance of norank_f__Muribaculaceae, and impact liver metabolic profiles, leading to significant amelioration of liver injury. The identification of Acetyl-DL-Valine as a protective metabolite underscores the therapeutic potential of targeting gut‒liver axis interactions in liver diseases. Conclusion: Overall, our data demonstrate that MSC-EVs exhibit nanotherapeutic potential in Con A-induced hepatitis and provide new insights into the treatment of autoimmune hepatitis. [ABSTRACT FROM AUTHOR]

Simple Summary: Rap1 has been implicated in the migration regulation of mesenchymal stem cells (MSCs). We found both Rap1A and Rap1B mRNAs expressed in human MSCs and suppressed their expression using siRNAs against both isoforms. Transwell migration, shear stress-dependent interaction, and adhesion to endothelial cells were all reduced in MSCs after the downregulation of Rap1A and/or Rap1B. In contrast, in a short-term homing model in immune-suppressed mice, MSCs treated with Rap1A+B siRNA showed increased accumulation in blood, liver, and spleen. Our data suggest that the modulation of Rap1A and/or B interferes with the adhesion, migration, and in vivo circulation behavior of human MSCs. Intravenously transplanted mesenchymal stromal cells (MSCs) have been shown to interact with endothelial cells and to migrate to tissues. However, intracellular signals regulating MSC migration are still incompletely understood. Here, we analyzed the role of Rap1 GTPase in the migration of human bone marrow-derived MSCs in vitro and in short-term homing in mice in vivo. MSCs expressed both Rap1A and Rap1B mRNAs, which were downregulated after treatment with siRNA against Rap1A and/or B. In a flow chamber model with pre-established human umbilical vein endothelial cells (HUVECs), Rap1A/B downregulated MSCs interacted for longer distances before arrest, indicating adhesion defects. CXCL12-induced adhesion of MSCs on immobilized Vascular Cell Adhesion Molecule (VCAM)-1 was also decreased after the downregulation of Rap1A, Rap1B, or both, as was CXCL12-induced transwell migration. In a competitive murine short-term homing model with i.v. co-injection of Rap1A+B siRNA-treated and control MSCs that were labeled with PKH 26 and PKH 67 fluorescent dyes, the Rap1A+B siRNA-treated MSCs were detected at increased frequencies in blood, liver, and spleen compared to control MSCs. Thus, Rap1 GTPase modulates the adhesion and migration of MSCs in vitro and may increase the bio-availability of i.v.-transplanted MSCs in tissues in a murine model. [ABSTRACT FROM AUTHOR]

Background: Renal cell cancer (RCC) is the most common and highly malignant subtype of kidney cancer. Mesenchymal stromal cells (MSCs) are components of tumor microenvironment (TME) that influence RCC progression. The impact of RCC-secreted small non-coding RNAs (sncRNAs) on TME is largely underexplored. Here, we comprehensively analysed the composition of exosomal sncRNAs secreted by RCC cells to identify those that influence MSCs. Methods: Exosomal sncRNAs secreted by RCC cells and normal kidney cells were analyzed using RNAseq, followed by qPCR validation. MSCs were treated by conditioned media (CM) derived from RCC cells and transfected with piRNA, followed by the analysis of proliferation, viability, migration and immunocytochemical detection of piRNA. Expression of MSCs genes was evaluated using microarray and qPCR. TCGA data were analyzed to explore the expression of sncRNAs in RCC tumors. Results: RNAseq revealed 40 miRNAs, 71 tRNAs and four piRNAs that were consistently secreted by RCC cells. qPCR validation using five independent RCC cell lines confirmed that expressions of miR-10b-3p and miR-125a-5p were suppressed, while miR-365b-3p was upregulated in exosomes from RCC cells when compared with normal kidney proximal tubules. The expression of miR-10b-3p and miR-125a-5p was decreased, whereas the expression of miR-365b-3p was increased in RCC tumors and correlated with poor survival of patients. Expressions of tRNA-Glu, tRNA-Gly, and tRNA-Val were the most increased, while tRNA-Gln, tRNA-Leu, and tRNA-Lys were top decreased in RCC exosomes when compared with normal kidney cells. Moreover, hsa_piR_004153, hsa_piR_016735, hsa_piR_019521, and hsa_piR_020365 were consistently upregulated in RCC exosomes. piR_004153 (DQ575660.1; aliases: hsa_piRNA_18299, piR-43772, piR-hsa-5938) was the most highly expressed in exosomes from RCC cells when compared with normal kidney cells. Treatment of MSCs with RCC CM resulted in upregulation of piR_004153 expression. Transfection of MSCs with piR_004153 stimulated their migration and viability, and altered expression of 35 genes, including downregulation of FGF2, SLC7A5, and WISP1. Immunocytochemistry confirmed the nuclear localization of piR_004153 transfected in MSCs. Conclusion: RCC cells secrete multiple sncRNAs, including piR_004153 which targets MSCs, alters expression of FGF2, SLC7A5, and WISP1, and stimulates their motility and viability. To our knowledge, this is the first study showing that cancer-derived piRNA can enhance MSC migration. [ABSTRACT FROM AUTHOR]

Purpose: Preculturing isolated islets with Mesenchymal Stromal Cells (MSCs) improves their functional survival in vitro and subsequent transplantation outcomes in vivo. The MSC secretory product Annexin A1 (ANXA1) is a key modulator of MSC-mediated improvements in islet function. The current study aims to determine the influence of MSCs and defined MSC secretory products, including ANXA1, on the inflammatory crosstalk between isolated islets and Endothelial Cells (ECs), using in vitro models of the clinically-preferred intraportal islet transplantation niche. Methods: Islets were cultured alone, with MSCs, or with MSC secretory products and exposed to pro-inflammatory cytokines. Islet gene expression of C-C Motif Chemokine Ligand 2 (CCL2), C-X-C Motif Chemokine Ligand (CXCL)-10 (CXCL10) and CXCL1 were assessed by RT-qPCR. EC activation was induced with 100 U/ml TNF for 24 h. Islet-EC co-cultures were used to determine the influence of MSCs, or MSC secretory products on the inflammatory crosstalk between isolated islets and ECs. VCAM-1 and ICAM-1 expression were assessed at the mRNA and protein level in ECs, using RT-qPCR and immunofluorescence. Results: MSCs reduce pro-inflammatory cytokine-induced islet CCL2, CXCL10, and CXCL1 gene expression, which is partially mimicked by ANXA1. MSCs and ANXA1 have a similar capacity to reduce TNF-induced EC activation. Isolated islets exacerbate TNF-induced EC activation. Preculturing islets with MSCs reduces islet-exacerbated EC activation. ANXA1 reduces islet-exacerbated EC activation, when present during the islet preculture and islet-EC co-culture period. Conclusion: MSC-derived secretory factors, including ANXA1, may be used in islet transplantation protocols to target donor islet and host EC inflammation at the intraportal niche. Highlights: MSCs and MSC-derived secretory factors, including Annexin A1, reduce EC activation. MSCs and MSC-secretory factors reduce islet expression of pro-inflammatory chemokines. MSCs and MSC-secretory factors reduce inflammatory crosstalk between islets and ECs. [ABSTRACT FROM AUTHOR]

Although elderly osteoporotic patients have similar implant survival rates compared with those of normal individuals, they require longer healing periods to achieve proper osseointegration. This may be related to chronic inflammatory responses and impaired stem cell repair functions in the osteoporotic bone microenvironment. Recently, the deubiquitinating enzyme, ubiquitin-specific peptidase 7 (USP7), was found to regulate the macrophage immune response and modulate stem cell osteogenic differentiation. The selective inhibitor of USP7, P5091, has also been found to promote bone repair and homeostasis in osteoporotic conditions. However, the roles of USP7 and P5091 in osteoimmunology and dental implant osseointegration under senile osteoporotic conditions remain unclear. In this study, USP7 depletion and P5091 were shown to inhibit inflammation in senescent bone marrow–derived macrophages (BMDMs) and promote osteogenic differentiation in aged bone marrow mesenchymal stromal cells (BMSCs). Furthermore, mRNA-Seq revealed that USP7 depletion could enhance efferocytosis in senescent BMDMs through the EPSIN1/low-density lipoprotein receptor-related protein 1 (LRP1) pathway and selectively induce apoptosis (senolysis) in aged BMSCs. In senile osteoporotic mice, we found that the osseointegration period was prolonged compared with young mice, and P5091 promoted the early stage of osseointegration, which may be related to macrophage efferocytosis around the implant. Collectively, this study suggests that USP7 inhibition may accelerate the osseointegration process in senile osteoporotic conditions by promoting macrophage efferocytosis and aged BMSCs apoptosis. This has implications for understanding the cellular interactions and signaling mechanisms in the peri-implant bone microenvironment under osteoporotic conditions. It may also provide clinical significance in developing new therapies to enhance osseointegration quality and shorten the edentulous period in elderly osteoporotic patients. [ABSTRACT FROM AUTHOR]

Introduction: Systemic sclerosis (SSc) is the rheumatic disease with the highest individual mortality rate with a detrimental impact on quality of life. Cell-based therapies may offer new perspectives for this disease as recent phase I trials support the safety of IV infusion of allogeneic mesenchymal stromal cells in SSc and case reports highlight the potential use of Chimeric Antigen Receptor (CAR)-T cells targeting CD19 in active SSc patients who have not responded to conventional immunosuppressive therapies. Areas covered: This narrative review highlights the most recent evidence supporting the use of cellular therapies in SSc as well as their potential mechanisms of action and discusses future perspectives for cell-based therapies in SSc. Medline/PubMed was used to identify the articles of interest, using the keywords 'Cellular therapies,' 'Mesenchymal stromal cells,' 'Chimeric Antigen Receptor' AND 'systemic sclerosis.' Milestones articles reported by the authors were also used. Expert opinion: Cellular therapies may represent an opportunity for long-term remission/cure in patients with different autoimmune diseases, including SSc who have not responded to conventional therapies. Multiple ongoing phase I/II trials will provide greater insights into the efficacy and toxicity of cellular therapies. [ABSTRACT FROM AUTHOR]

Background: Hemorrhagic shock (HS) corresponds to absolute hypovolemia creating an imbalance between oxygen supply and consumption. This causes an impaired hemostasis, a systemic inflammatory response, and microvascular permeability which can lead to multiple organ failure (MOF). There is no specific treatment for the endothelial dysfunction that plays a major role in the evolution towards MOF. Mesenchymal stromal cells (MSC) have been used in clinical trials for their immunomodulation and tissue repair capabilities for many years. Moreover, we previously showed that IL-1β-primed-MSC (MSCp) attenuated HS-induced organ injuries. The objective of the study was to determine whether MSCp could prevent the onset of MOF after HS by preventing endothelial dysfunction. Methods: We established a rat model of HS, inducing 90 min of HS at a fixed mean arterial pressure of 35 mmHg, followed by resuscitation and transfusion. MSCp treatment was administered intravenously at the onset of resuscitation. After 6 h, we assessed plasma levels of endothelial markers, vascular permeability using Evans Blue (EB) dye, and renal and hepatic water content by measuring the wet-to-dry weight difference. Additionally, we investigated the ability of MSCp to inhibit leukocyte adhesion to activated endothelium in vitro. Results: Our results indicate that early administration of MSCp significantly reduced the percentage of water content and EB dye in the liver but not in the kidney. These results were associated with a trend toward decreased plasma levels of Syndecan-1, ICAM-1, vWF, and VCAM-1. In vitro, MSCp reduced leukocyte-endothelial cell adhesion. Together, our results suggest that MSCp help to prevent endothelial dysfunction and vascular leakage, which, in turn, could protect the liver from injury. [ABSTRACT FROM AUTHOR]

Constructing artificial tertiary lymphoid structures (TLSs) opens new avenues for advancing cancer immunotherapy and personalized medicine by creating controllable immune niches. Mesenchymal stromal cells (MSCs) offer an ideal stromal source for such constructs, given their potent immunomodulatory abilities and accessibility. In this study, we explored the potential of adipose-derived MSCs to adopt TLS-supportive phenotypes and facilitate lymphocyte organization. Single-cell RNA sequencing revealed a distinct subpopulation of MSCs expressing key fibroblastic reticular cell (FRC)-associated markers, including IL-7, PDPN, and IL-15, though lacking follicular dendritic cell (FDC) markers. TNF-α stimulation, but not LTα2β1, further enhanced FRC marker expression (IL-7, PDPN, and ICAM1). Notably, in 3D spheroid co-culture with lymphocytes, MSCs upregulated additional FRC markers, specifically CCL21. Upon implantation into adipose tissue, MSC-lymphocyte organoids maintained structural integrity and showed extensive T-cell infiltration and partial vascularization after 15 days in vivo, although organized B-cell follicles and FDC markers were still lacking. These findings highlight MSCs' intrinsic ability to adopt an FRC-like phenotype that supports T-cell and HEV organization, suggesting that further optimization, including genetic modification, may be needed to achieve an FDC phenotype and replicate the full architectural and functional complexity of TLSs. [ABSTRACT FROM AUTHOR]

The present study investigates the influence of nitrosamines and etoposide on mesenchymal stromal cells (MSCs) in a differentiation state- and biological age-dependent manner. The genotoxic effects of the agents on both neonatal and adult stem cell populations after treatment, before, or during the course of differentiation, and the sensitivity of the different MSC types to different concentrations of MNU or etoposide were assessed. Hereby, the multipotent differentiation capacity of MSCs into osteoblasts, adipocytes, and chondrocytes was analyzed. Our findings reveal that while all cell types exhibit DNA damage upon exposure, neonatal CB-USSCs demonstrate enhanced resistance to genotoxic damage compared with their adult counterparts. Moreover, the osteogenic differentiation of MSCs was more susceptible to genotoxic damage, whereas the adipogenic and chondrogenic differentiation potentials did not show any significant changes upon treatment with genotoxin. Furthermore, we emphasize the cell-specific variability in responses to genotoxic damage and the differences in sensitivity and reaction across different cell types, thus advocating the consideration of these variabilities during drug testing and developmental biological research. [ABSTRACT FROM AUTHOR]

Background: Mesenchymal stromal cells (MSC) have immunomodulatory and hematopoiesis-supporting properties that could potentially benefit hematopoietic stem cell (HSC) engraftment and decrease the incidence and/or severity of graft-versus-host disease (GVHD). Methods: Based on our previous pilot study, we established a multicenter, prospective, randomized, double-blind trial evaluating the efficacy of co-infusing third-party MSC (1.5–3 × 106/kg) versus placebo on the day of HSC transplantation (HCT) to prevent GVHD in recipients of HLA-mismatched unrelated donors after reduced-intensity conditioning. Results: The study planned to include 120 patients to improve 1-year overall survival (OS) from 55 to 77% but was stopped after 9 years for low recruitment (n = 38). One-year OS was 74% in the MSC group and 80% in the placebo group. In multivariate analysis, the incidence of grade II-IV acute GVHD was significantly lower in patients receiving MSC (HR 0.332, 95% CI 0.124–0.890, p = 0.0284). No difference was observed in the incidences of chronic GVHD, infection or relapse, overall or progression-free survival at 1 year or long-term, or hematopoietic and immune reconstitution. Conclusions: Despite premature study closure, the suggested beneficial effect of MSC co-transplantation for the prevention of acute GVHD in HLA-mismatched HCT warrants further investigation. [ABSTRACT FROM AUTHOR]

Erectile dysfunction (ED) is a major threat to male fertility and quality of life, and mesenchymal stromal cells (MSCs) are a promising therapeutic option. However, therapeutic outcomes are compromised by low MSC retention and survival rates in corpus cavernosum tissue. Here, we developed an innovative magnetic soft microrobot comprising an ultrasoft hydrogel microsphere embedded with a magnetic nanoparticle chain for MSC delivery. This design also features phenylboronic acid groups for scavenging reactive oxygen species (ROS). With a Young's modulus of less than 1 kPa, the ultrasoft microrobot adapts its shape within narrow blood vessels, ensuring a uniform distribution of MSCs within the corpus cavernosum. Our findings showed that compared with traditional MSC injections, the MSC delivery microrobot (MSC-Rob) significantly enhanced MSC retention and survival. In both rat and beagle ED models, MSC-Rob treatment accelerated the repair of corpus cavernosum tissue and restored erectile function. Single-cell RNA sequencing (scRNA-seq) revealed that MSC-Rob treatment facilitates nerve and blood vessel regeneration in the corpus cavernosum by increasing the presence of regenerative macrophages. Overall, our MSC-Rob not only advances the clinical application of MSCs for ED therapy but also broadens the scope of microrobots for other cell therapies. [ABSTRACT FROM AUTHOR]

Background: The ability of mesenchymal stromal cells (MSCs) to facilitate regenerative responses in inflamed and injured tissues, coupled with preclinical data suggesting potential to restore defective collagen VII at the dermo-epidermal junction, has raised the hope that MSCs may provide an effective disease-modifying therapy for patients suffering from recessive dystrophic epidermolysis bullosa (RDEB). Methods: We present a descriptive analysis of the clinical research on systemic MSC administration to RDEB patients available in PubMed, including six early-phase studies and one case report, involving 59 patients who received 1–3 intravenous infusions of MSCs from various sources. Results: Based on 133 MSC infusions, a total of 44 mostly mild adverse events were reported as definitely, possibly or likely related to the study treatment, only two of which led to treatment discontinuation. Improvements were seen in skin manifestations, disease activity, pain, pruritus and quality of life, with considerable heterogeneity in reported outcome variables and measurement tools between studies, and large inter-patient variability within studies. Conclusions: Although the current evidence base is limited, reflecting the typical challenges of clinical research in rare diseases, the reported results suggest potential treatment benefits for patients and provide a rationale for continuing to pursue this therapeutic approach. [ABSTRACT FROM AUTHOR]

Inflammatory bowel disease (IBD) is characterized by periods of flare-ups and remission. It is likely to be an autoimmune in origin, presenting persistent therapeutic challenges despite current therapies. This study aims to investigate the potential of umbilical cord mesenchymal stromal cells (UCMSCs) in treating ulcerative colitis (UC). This study is a prospective phase 1 pilot, open-label, single-arm, and single-center study. UCMSCs were cultured under current Good Manufacturing Practice (cGMP) conditions and intravenously administered to six patients with UC. Safety and efficacy were evaluated using the Mayo Score/Disease Activity Index. Among the six enrolled adult patients, five completed long-term follow-ups. All exhibited at diagnosis active UC confirmed through comprehensive assessment methods. Each patient received three injections intravenously 2 weeks apart with a dose of 100 million UCMSC each. No significant short-term or intermediate-term adverse events were detected post-UCMSC administration. Long-term follow-up at 12 and 24 months showed sustained safety and no adverse events. Notably, three out of five patients achieved a Mayo score of 0 for UC, maintained at both 12 and 24 months, indicating a highly significant response (P < 0.001). This study demonstrates the safety and potential efficacy of UCMSCs in active UC. However, larger trials are warranted to validate these preliminary findings and to establish the role of UCMSC therapy as an option for managing UC. [ABSTRACT FROM AUTHOR]

Abnormal mechanical loading is one of the major risk factors for articular cartilage degeneration. Engineered mesenchymal stromal cell (MSC)‐derived cartilage holds great promise for cell‐based cartilage repair. However, physiological loading protocols were shown to reduce matrix synthesis of MSC‐derived neocartilage in vitro and the regulators of this undesired mechanoresponse remain poorly understood. Parathyroid hormone‐related protein (PTHrP) is involved in cartilage development and can affect extracellular matrix (ECM) production during MSC chondrogenesis opposingly, depending on a continuous or transient exposure. PTHrP is induced by various mechanical cues in multiple tissues and species; but whether PTHrP is regulated in response to loading of human engineered neocartilage and may affect matrix synthesis in a positive or negative manner is unknown. The aim of this study was to investigate whether dynamic loading adjusts PTHrP‐signaling in human MSC‐derived neocartilage and whether it regulates matrix synthesis and other factors involved in the MSC mechanoresponse. Interestingly, MSC‐derived chondrocytes significantly upregulated PTHrP mRNA (PTHLH) expression along with its second messenger cAMP in response to loading in our custom‐built bioreactor. Exogenous PTHrP(1‐34) induced the expression of known mechanoresponse genes (FOS, FOSB, BMP6) and significantly decreased glycosaminoglycan (GAG) and collagen synthesis similar to loading. The adenylate‐cyclase inhibitor MDL‐12,330A rescued the load‐mediated decrease in GAG synthesis, indicating a direct involvement of cAMP‐signaling in the reduction of ECM production. According to COL2A1‐corrected hypertrophy‐associated marker expression, load and PTHrP treatment shared the ability to reduce expression of MEF2C and PTH1R. In conclusion, the data demonstrate a significant mechanoinduction of PTHLH and a negative contribution of the PTHrP‐cAMP signaling axis to GAG synthesis in MSC‐derived chondrocytes after loading. To improve ECM synthesis and the mechanocompetence of load‐exposed neocartilage, inhibition of PTHrP activity should be considered for MSC‐based cartilage regeneration strategies. [ABSTRACT FROM AUTHOR]

Background/Objectives: Cartilage injuries and osteoarthritis are prevalent public health problems, due to their disabling nature and economic impact. Mesenchymal stromal cells (MSCs) isolated from different tissues have the immunomodulatory capacity to regulate local joint environment. This translational study aims to compare cartilage restoration from MSCs from the synovial membrane (SM) and dental pulp (DP) by a tissue-engineered construct with Good Manufacturing Practices. Methods: A controlled experimental study was conducted on fourteen miniature pigs, using scaffold-free Tissue Engineering Constructs (TECs) from DP and SM MSCs, with a 6-month follow-up. Total thickness cartilage defects were created in both hind knees; one side was left untreated and the other received a TEC from either DP (n = 7) or SM (n = 7). An MRI assessed the morphology using the MOCART scoring system, T2 mapping evaluated water, and collagen fiber composition, and histological analysis was performed using the ICRS-2 score. Results: The untreated group had a mean MOCART value of 46.2 ± 13.4, while the SM-treated group was 65.7 ± 15.5 (p < 0.05) and the DP-treated group was 59.0 ± 7.9 (n.s.). The T2 mapping indicated a mean value of T2 of 54.9 ± 1.9 for native cartilage, with the untreated group at 50.9 ± 2.4 (p < 0.05). No difference was found between the T2 value of native cartilage and the treated groups. The ICRS-2 mean values were 42.1 ± 14.8 for the untreated group, 64.3 ± 19.0 for SM (p < 0.05), and 54.3 ± 12.2 for DP (n.s.). Conclusion: MRI and histological analysis indicated that TEC treatment led to superior cartilage coverage and quality compared to the defect group. TECs from SM demonstrated better results than the defect group in the histological assessment. [ABSTRACT FROM AUTHOR]

Cartilage degeneration is a key feature of aging and osteoarthritis, characterized by the progressive deterioration of joint function, pain, and limited mobility. Current treatments focus on symptom relief, not cartilage regeneration. Mesenchymal stromal cells (MSCs) offer a promising therapeutic option due to their capability to differentiate into chondrocytes, modulate inflammation, and promote tissue regeneration. This review explores the potential of MSCs for cartilage regeneration, examining their biological properties, action mechanisms, and applications in preclinical and clinical settings. MSCs derived from bone marrow, adipose tissue, and other sources can self-renew and differentiate into multiple cell types. In aging cartilage, they aid in tissue regeneration by secreting growth factors and cytokines that enhance repair and modulate immune responses. Recent preclinical studies show that MSCs can restore cartilage integrity, reduce inflammation, and improve joint function, although clinical translation remains challenging due to limitations such as cell viability, scalability, and regulatory concerns. Advancements in MSC delivery, including scaffold-based approaches and engineered exosomes, may improve therapeutic effectiveness. Potential risks, such as tumorigenicity and immune rejection, are also discussed, emphasizing the need for optimized treatment protocols and large-scale clinical trials to develop effective, minimally invasive therapies for cartilage regeneration. [ABSTRACT FROM AUTHOR]

Although adipose stem cell (ASC)-derived extracellular vesicles (EVs) are as effective as ASCs in the suppression of Th2 cell-mediated eosinophilic inflammation, the role of identified pulmonary genes has not been well documented. Thus, we assessed the immunomodulatory effects of paraoxonase-1 (PON1) on allergic airway inflammation in a mouse model of asthma. Five-week-old female C57BL/6 mice were sensitized to ovalbumin (OVA) by intraperitoneal injection and challenged intranasally with OVA. To evaluate the effect of PON1 on allergic airway inflammation, the intranasal and intraperitoneal injections of recombinant mouse serum PON1 (5 μg/50 μL) were performed before the OVA challenge. We evaluated airway hyperresponsiveness (AHR), total inflammatory cells, and eosinophils in the bronchoalveolar lavage fluid (BALF), lung histology, serum immunoglobulin (Ig), cytokine profiles of BALF and lung draining lymph nodes (LLNs), the expression of interleukin (IL)-25 and transforming growth factor (TGF)-β in mouse lung epithelial cell (MLE-12 cell), and dendritic cell (DC) differentiation. The intraperitoneal and intranasal administration of PON1 significantly decreased AHR, total inflammatory cells and eosinophils in BALF, eosinophilic airway inflammation, serum total, and OVA-specific IgE. PON1 treatment, which marked reduced IL-4, IL-5, and IL-13 in the BALF and LLN but significantly increased interferon-γ and TGF-β. Furthermore, PON1 treatment significantly decreased the expression of IL-25 and increased TGF-β in MLE-12 cells. The expressions of CD40, CD80, and CD86 in immature DCs were significantly increased by PON1 treatment. The administration of PON1 ameliorated allergic airway inflammation and improved AHR through the downregulation of IL-4, IL-5, and IL-13 and upregulation of TGF-β in asthmatic mice. Furthermore, PON1 treatment decreased Th2-mediated inflammation induced by Aspergillus protease antigen by decreasing IL-25 and increasing TGF-β. [ABSTRACT FROM AUTHOR]