Ability Of Mesenchymal Stem Cells Research Articles

KDM2B and its peptides promote the stem cells from apical papilla mediated nerve injury repair in rats by intervening EZH2 function.

How to improve the neurogenic potential of mesenchymal stem cells (MSCs) and develop biological agent based on the underlying epigenetic mechanism remains a challenge. Here, we investigated the effect of histone demethylase Lysine (K)-specific demethylase 2B (KDM2B) on neurogenic differentiation and nerve injury repair by using MSCs from dental apical papilla (SCAP). We found that KDM2B promoted the neurogenic indicators expression and neural spheres formation in SCAP, and modified the Histone H3K4 trimethylation (H3K4me3) methylation on neurogenesis-related genes. KDM2B improved the SCAP mediated recovery of motor ability at the early healing stage of spinal cord injury rats. Meanwhile, KDM2B acted as a negative regulator to its partner EZH2 during neurogenic differentiation, enhancer of zeste homologue 2 (EZH2) suppressed the neurogenic ability of SCAP. Further, the protein interaction between KDM2B and EZH2 was identified which decreased during neurogenic differentiation. On this basis, we revealed seven key protein binding sequences of KDM2B to EZH2, and synthesized KDM2B-peptides based on these sequences. By the usage of KDM2B-peptides, EZH2 function was effectively intervened and the neurogenic ability of SCAP was promoted. More, KDM2B-peptides significantly improved the SCAP mediated functional recovery at SCI early phase. Our study revealed that KDM2B acted as a promotor to neurogenic differentiation ability of dental MSCs through binding and negatively regulating EZH2, and provided the KDM2B-peptides as candidate agents for improving the neurogenic ability of MSCs and nerve injury repair.

The impact of 980nm diode laser irradiation on the proliferation of mesenchymal stem cells derived from the umbilical cord's

Various cell types can have their growth accelerated by using low-intensity laser radiation. The study is intended to look at the impacts of laser radiation at low energy intensity on the ability of mesenchymal stem cells (MSCs) generated from umbilical cords to proliferate as well as survive in low-nutrient conditions. The study applied two different energy densities, 2.5 j/cm2 and 5 j/cm2, using a 980 nm diode laser radiation. This allowed for the observation of the effects of these specific elements on the behavior of the cells in a controlled environment at various concentrations of fetal bovine (7.5 %, 10 %, 12.5 %, and 15 %). The cells were grown in a medium lacking in nutrients and were enriched with varying quantities of serum from fetal bovines. The MTT test was used to evaluate the mitochondrial activity of the cell. Following 72 hours, it was shown that cells treated with 2.5 j/cm2 and 10 % fetal bovine serum had significantly higher MTT test activity than cells treated with 5 j/cm2.The results of this study show that even in the presence of dietary deficiencies, low-intensity laser radiation therapy can stimulate the growth of mesenchymal stem cells isolated from umbilical cords.

Mesenchymal Stem Cells and Secretome as a New Possible Approach to Treat Cartilage Damage: An In Vitro Study.

Introduction: Osteoarthritis is a degenerative condition of the cartilage, often common among the population and occurs frequently with aging. Many factors are decisive for the development of its pathogenesis such as age, obesity, trauma, mechanical load, and modification of synovial biology. The main features of osteoarthritis are chondrocytes and cartilage matrix loss, which lead to pain, loss of function of the whole joint, and disability, representing a relevant health problem. Recently, a new therapeutic approach based on cell therapy has been studying the regenerative ability of mesenchymal stem cells for osteoarthritic chondrocytes. Aim: This in vitro study clarifies the regenerative effects of multipotent adipose-derived stem cells and the pluripotent amniotic epithelial stem cells on arthrosis chondrocytes by performing co-culture experiments. Methods: We studied the regenerative potential of secretome (soluble factors and extracellular vesicles), mesenchymal stem cells, and the adipose stromal vascular fraction. The regenerative effects were evaluated by gene and protein expression analysis of articular cartilage-specific genes and proteins like col2a1, acan, and sox9. Results: Mesenchymal stem cells, secretome, and adipose stromal vascular fractions influenced the cartilage genes and protein expression. Conclusions: The results indicate that the treatment with mesenchymal stem cells could be the best biological approach for cartilage regenerative medicine.

Efficiently directing differentiation and homing of mesenchymal stem cells to boost cartilage repair in osteoarthritis via a nanoparticle and peptide dual-engineering strategy

Mesenchymal stem cells (MSCs) are expected to be useful therapeutics in osteoarthritis (OA), the most common joint disorder characterized by cartilage degradation. However, evidence is limited with regard to cartilage repair in clinical trials because of the uncontrolled differentiation and weak cartilage-targeting ability of MSCs after injection. To overcome these drawbacks, here we synthesized CuO@MSN nanoparticles (NPs) to deliver Sox9 plasmid DNA (favoring chondrogenesis) and recombinant protein Bmp7 (inhibiting hypertrophy). After taking up CuO@MSN/Sox9/Bmp7 (CSB NPs), the expressions of chondrogenic markers were enhanced while hypertrophic markers were decreased in response to these CSB-engineered MSCs. Moreover, a cartilage-targeted peptide (designated as peptide W) was conjugated onto the surface of MSCs via a click chemistry reaction, thereby prolonging the residence time of MSCs in both the knee joint cavity of mice and human-derived cartilage. In a surgery-induced OA mouse model, the NP and peptide dual-modified W-CSB-MSCs showed an enhancing therapeutic effect on cartilage repair in knee joints compared with other engineered MSCs after intra-articular injection. Most importantly, W-CSB-MSCs accelerated cartilage regeneration in damaged cartilage explants derived from OA patients. Thus, this new peptide and NPs dual engineering strategy shows potential for clinical applications to boost cartilage repair in OA using MSC therapy.

Pharmacological activation of mesenchymal stem cells increases gene expression pattern of cell adhesion molecules and fusion with neonatal cardiomyocytes.

Cellular therapy is considered a better option for the treatment of degenerative disorders. Different cell types are being used for tissue regeneration. Despite extensive research in this field, several issues remain to be addressed concerning cell transplantation. One of these issues is the survival and homing of administered cells in the injured tissue, which depends on the ability of these cells to adhere. To enhance cell adherence and survival, Rap1 GTPase was activated in mesenchymal stem cells (MSCs) as well as in cardiomyocytes (CMs) by using 8-pCPT-2'-O-Me-cAMP, and the effect on gene expression dynamics was determined through quantitative reverse transcriptase-polymerase chain reactionanalysis. Pharmacological activation of MSCs and CMsresulted in the upregulation of connexin-43 and cell adhesion genes, which increased the cell adhesion ability of MSCs and CMs, and increased the fusion of MSCs with neonatal CMs. Treating stem cells with a pharmacological agent that activates Rap1a before transplantation can enhance their fusion with CMs and increase cellular regeneration.

Osteostaticytes: A novel osteoclast subset couples bone resorption and bone formation

BackgroundOsteomyelitis (OM) is an inflammatory condition of bone characterized by cortical bone devascularization and necrosis. Dysregulation of bone remodelling is triggered by OM. Bone remodelling is precisely coordinated by bone resorption and formation via a reversal phase. However, the cellular and molecular mechanisms underlying bone remodelling failure after osteomyelitis remain elusive. MethodsTo elucidate the cellular and molecular mechanism underlying bone healing after osteomyelitis, we employed single-cell RNA sequencing (scRNA-seq) to depict the atlas of human cortical bone in normal, infected and reconstructed states. Dimensionality reduction by t-stochastic neighbourhood embedding (t-SNE) and graph-based clustering were applied to analyse the detailed clusters of osteoclast lineages. After trajectory analysis of osteoclast lineages over pseudotime, real-time PCR and immunofluorescence (IF) staining were applied to identify marker gene expression of various osteoclast lineages in the osteoclast induction model and human bone sections, respectively. The potential function and communication of osteoclasts were analysed via gene set enrichment analysis (GSEA) and CellChat. The chemotactic ability of mesenchymal stem cells (MSCs) and osteoclast lineage cells in various differentiation states was determined by transwell assays and coculture assays. The effects of various osteoclast lineages on the osteogenic differentiation potential of MSCs were also determined by using this coculture system. A normal mouse tibia fracture model and an osteomyelitis-related tibia fracture model were generated via injection of luciferase-labelled Staphylococcus aureus to verify the relationships between a novel osteoclast lineage and MSCs. Then, the infection was detected by a bioluminescence imaging system. Finally, immunofluorescence staining was used to detect the expression of markers of MSCs and novel osteoclast lineages in different remodelling phases in normal and infected bone remodelling models. ResultsIn this study, we constructed a cell atlas encompassing normal, infected, and reconstructed cortical bone. Then, we identified a novel subset at the earlier stage of the osteoclast lineage that exhibited increased expression of IDO1, CCL3, and CCL4. These IDO1highCCL3highCCL4high cells, termed osteostaticytes (OSCs), were further regarded as the reservoir of osteoclasts in the reversal phase. Notably, OSCs exhibited the highest chemotactic activity, surpassing other lineage subsets. We also discovered that cells at the earlier stage of the osteoclast lineage play a significant role in recruiting mesenchymal stem cells (MSCs). Finally, the data revealed that OSCs might be positively related to the occurrence of bone MSCs and the contribution of bone remodelling. ConclusionCollectively, our findings revealed a novel stage (OSC) within the osteoclast lineage, potentially representing elusive bone reversal cells due to its increased chemotactic ability towards MSCs and potential contribution to bone remodelling. This study provides valuable insights into the intricate mechanisms of the reversal phase during bone remodelling and unveils potential therapeutic strategies for diseases associated with bone uncoupling. Translational potential of this articleThis study identified a new subset, referred to as IDO1(plus symbol) CCL3(plus symbol) CCL4(plus symbol) osteostaticytes which displayed the highest chemotactic activity among all osteoclast lineages and may serve as reversal cells in bone remodelling. These findings offer new insights and insights for understanding bone reversal-related diseases and may serve as novel therapeutic targets for conditions such as osteomyelitis and delayed bone healing.

3D modular bioceramic scaffolds for the investigation of the interaction between osteosarcoma cells and MSCs

Recent advances in bone tissue engineering have shown promise for bone repair post osteosarcoma excision. However, conflicting research on mesenchymal stem cells (MSCs) has raised concerns about their potential to either promote or inhibit tumor cell proliferation. It is necessary to thoroughly understand the interactions between MSCs and tumor cells. Most previous studies only focused on the interactions between cells within the tumor tissues. It has been challenging to develop an in vitro model of osteosarcoma excision sites replicating the complexity of the bone microenvironment and cell distribution. In this work, we designed and fabricated modular bioceramic scaffolds to assemble into a co-culture model. Because of the bone-like composition and mechanical property, tricalcium phosphate bioceramic could mimic the bone microenvironment and recapitulate the cell-extracellular matrix interaction. Moreover, the properties for easy assembly enabled the modular units to mimic the spatial distribution of cells in the osteosarcoma excision site. Under this co-culture model, MSCs showed a noticeable tumor-stimulating effect with a potential risk of tumor recurrence. In addition, tumor cells also could inhibit the osteogenic ability of MSCs. To undermine the stimulating effects of MSCs on tumor cells, we present the methods of pre-differentiated MSCs, which had lower expression of IL-8 and higher expression of osteogenic proteins. Both in vitro and in vivo studies confirm that pre-differentiated MSCs could maintain high osteogenic capacity without promoting tumor growth, offering a promising approach for MSCs’ application in bone regeneration. Overall, 3D modular scaffolds provide a valuable tool for constructing hard tissue in vitro models. Statement of significanceBone tissue engineering using mesenchymal stem cells (MSCs) and biomaterials has shown promise for bone repair post osteosarcoma excision. However, conflicting researches on MSCs have raised concerns about their potential to either promote or inhibit tumor cell proliferation. It remains challenges to develop in vitro models to investigate cell interactions, especially of osteosarcoma with high hardness and special composition of bone tissue. In this work, modular bioceramic scaffolds were fabricated and assembled to co-culture models. The interactions between MSCs and MG-63 were manifested as tumor-stimulating and osteogenesis-inhibiting, which means potential risk of tumor recurrence. To undermine the stimulating effect, pre-differentiation method was proposed to maintain high osteogenic capacity without tumor-stimulating, offering a promising approach for MSCs’ application in bone regeneration.

Effect of Fibroblast Growth Factor 2 and Low-Level Laser Therapy on the Adhesion and Proliferation of Periodontal Ligament Stem Cells.

Introduction: The adhesion ability of mesenchymal stem cells can significantly affect their viability and is considered a prerequisite for cell therapy. The current study sought to evaluate the effect of fibroblast growth factor 2 (FGF2) and low-level laser therapy (LLLT), either individually or in conjunction, on the adhesion and proliferation of periodontal ligament stem cells (PDLSCs) when applied on the first day of cell seeding. Methods: The experimental groups of this study comprised a control group and different combinations of adjunctive FGF2 (50 ng/mL) and LLLT with an 808 nm diode laser in one (LLLT-1) or two sessions (LLLT-2) of irradiation. The proliferation and adhesion of cells were evaluated by using the methylthiazolyl tetrazolium (MTT) assay and 4',6-diamidino-2-phenylindole (DAPI) staining. All experiments were done in triplicates on the first, third, and fifth days after cell seeding. Two-way ANOVA and post hoc Tukey tests were used to analyze the data of the MTT assay. P<0.05 was considered statistically significant. Results: One-day post-culture, only significant differences were found between the control group and the FGF2 (P=0.04) and FGF2+LLLT-2 application (P=0.04) groups. After three days post-cell culture, only a significantly higher proliferation rate was found in the control group than in the FGF2 group (P=0.01). After five days, the control group and LLLT-2 groups showed significantly higher amounts of proliferation compared to the other groups (P<0.05). DAPI staining qualitatively confirmed the results of the MTT assay. Conclusion: The LLLT can be applied to PDLSCs on the day of seeding without causing a notable decrease in their viability and adhesion. Conversely, the administration of FGF2 should be restricted on the seeding day and postponed to subsequent days as it may have adverse effects on their adhesion and proliferation.

KLF9 regulates osteogenic differentiation of mesenchymal stem cells.

Osteoporosis is a progressive skeletal disease which is characterized by reduced bone mass and degradation of bone microstructure. Mesenchymal stem cells (MSCs) have the potential to inhibit osteoporosis since they are multipotent stem cells that can differentiate into multiple types of cells including osteoblasts. Hence the mechanism of osteogenic differentiation of MSCs deserves comprehensive study. Here we report that KLF9 is a novel regulator in osteogenic differentiation of MSCs. We observed that depletion of KLF9 can largely compromise the osteogenic differentiation ability of MSCs. In addition, we revealed that inhibition of the PI3K-Akt pathway could also affect osteogenic differentiation since KLF9 depletion inhibits PI3K expression. Finally, we discovered that KLF9 expression can be induced by dexamethasone which is an essential component in osteogenic induction medium. Taken together, our study provides new insights into the regulatory role of KLF9 in osteogenic differentiation of MSCs.

ELABELA-APJ Axis Enhances Mesenchymal Stem Cell Proliferation and Migration via the METTL3/PI3K/AKT Pathway.

Mesenchymal stem cells (MSCs) possess a strong therapeutic potential in regenerative medicine. ELABELA (ELA) is a 32 amino acid peptide that binds to the apelin peptide jejunum receptor (APJ) to regulate cell proliferation and migration. The aim of this study was to investigate the function of ELA vis-a-vis the MSC proliferation and migration, and further explore the underlying mechanism. We demonstrated that the exogenous supplement of ELA boosts the proliferation and migration ability of MSCs, alongside improved in vitro cell viability. These capabilities were rendered moot upon APJ knockdown. In addition, ELA (5-20 μM) was shown to upregulate the expression of METTL3 in a concentrationdependent pattern, a capacity which was suppressed by APJ reduction, whereas the downregulation of METTL3 expression blocked the beneficial effects induced by ELA. ELA was also observed to upregulate the phosphorylation level of AKT. This ELA-induced activation of the PI3K/AKT pathway, however, is inhibited with knockdown of METTL3. Our data indicate that ELA could act as a promoter of MSC proliferation and migration in vitro through the APJ receptor, something which might be attributed to the activation of the METTL3/PI3K/AKT signaling pathway. Therefore, ELA is a candidate for optimizing MSC-based cell therapy, while METTL3 is a potential target for its promoting action on MSCs.

Study on liver tissue derived-extracellular vesicles regulating the osteogenic differentiation ability of mesenchymal stem cells and promoting the healing of jaw bone defects

Objective: To explore the biological process of liver tissue-derived extracellular vesicle (LT-EV) in promoting osteogenic differentiation of mesenchymal stem cells and healing of jaw defects to provide a feasible treatment method for the clinical treatment of jaw bone defects. Methods: Enzymatic hydrolysis and differential centrifugation were used to extract LT-EV, scanning electron microscopy, Western blotting, and nanoparticle tracking analyzers were used to identify and characterize LT-EV, and further to explore the biological functions of LT-EV through proteomics and Kyoto Encyclopedia of Genes and Genomes. Flow cytometry was used to detect LT-EV plasma concentration and to calculate the plasma half-life of LT-EV. Small animal in vivo imaging system was used to detect the biological distribution of LT-EV 24 hours after injection. Six C57BL/6 mice were divided into control group and LT-EV group (3 mice in each group) by simple random sampling method. All mice underwent jaw bone defect surgery and tail vein injection every 7 days (the control group was injected with phosphoric buffer saline, LT-EV group was injected with LT-EV), micro-CT was used to evaluate the degree of mouse jaw bone healing 28 days after surgery, HE staining was used to analyze the multi-organ biosafety of LT-EV, and immunofluorescence staining was used to detect the jaw bone expression of osteogenic marker proteins in the defect area. Human jaw bone mesenchymal stem cells (hJBMSC) induced by osteogenic differentiation were treated with LT-EV (obtained from orthognathic surgery patients provided by the Department of Traumatology and Orthognathic Surgery of School of Stomatology of The Fourth Military Medical University resected normal jaw bone fragments), and the difference in osteogenic differentiation ability between the hJBMSC group and the control group (phosphate buffer saline treatment) was compared, and the in vitro bone differentiation promoting effect of LT-EV was verified through alkaline phosphatase (ALP) staining and real-time fluorescence quantitative PCR. Results: The yield of LT-EV was high, and proteomics and Kyoto Encyclopedia of Genes and Genomes showed that LT-EV contained a series of proteins that regulated cell biological functions. LT-EV injected into the tail vein could reach the mouse jaw bone defect area and promote the regeneration and repair of the jaw bone defect [the bone volume fractions of the LT-EV group and the control group were (36.06±4.20)% and (18.58±5.61)%, respectively; t=4.32, P=0.013], and had good biosafety. LT-EV could promote osteogenic differentiation of hJBMSC in vitro. Compared to the control group, ALP staining and osteogenic gene expression levels were significantly enhanced after osteogenic differentiation of hJBMSC (P<0.05). Conclusions: LT-EV exhibits a high yield, ease of acquisition, high biological safety, and excellent bone-promoting effects. It holds promise as a novel cell-free therapy strategy for regenerating craniofacial bone defects.

Mesenchymal Stem Cell Spheroids: A Promising Tool for Vascularized Tissue Regeneration.

Mesenchymal stem cells (MSCs) are undifferentiated cells that can differentiate into specific cell lineages when exposed to the right conditions. The ability of MSCs to differentiate into particular cells is considered very important in biological research and clinical applications. MSC spheroids are clusters of MSCs cultured in three dimensions, which play an important role in enhancing the proliferation and differentiation of MSCs. MSCs can also participate in vascular formation by differentiating into endothelial cells and secreting paracrine factors. Vascularization ability is essential in impaired tissue repair and function recovery. Therefore, the vascularization ability of MSCs, which enhances angiogenesis and accelerates tissue healing has made MSCs a promising tool for tissue regeneration. However, MSC spheroids are a relatively new research field, and more research is needed to understand their full potential. In this review, we highlight the importance of MSC spheroids' vascularization ability in tissue engineering and regenerative medicine while providing the current status of studies on the MSC spheroids' vascularization and suggesting potential future research directions for MSC spheroids. Studies both in vivo and in vitro have demonstrated MSC spheroids' capacity to develop into endothelial cells and stimulate vasculogenesis. MSC spheroids show potential to enhance vascularization ability in tissue regeneration. Yet, further research is required to comprehensively understand the relationship between MSC spheroids and vascularization mechanisms.

Establishment and Evaluation Strategy of an in vitro Cell Model of Bone Marrow Microenvironment Injury in Mouse Acute Graft-Versus-Host Disease

To establish a mesenchymal stem cell（MSC）-based in vitro cell model for the evaluation of mouse bone marrow acute graft-versus-host disease (aGVHD). Female C57BL/6N mice aged 6-8 weeks were used as bone marrow and lymphocyte donors, and female BALB/c mice aged 6-8 weeks were used as aGVHD recipients. The recipient mouse received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) in 6-8 hours post irradiation to establish a bone marrow transplantation (BMT) mouse model (n=20). In addition, the recipient mice received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) and spleen lymphocytes (2×106/mouse) in 6-8 hours post irradiation to establish a mouse aGVHD model (n=20). On the day 7 after modeling, the recipient mice were anesthetized and the blood was harvested post eyeball enucleation. The serum was collected by centrifugation. Mouse MSCs were isolated and cultured with the addition of 2%, 5%, and 10% recipient serum from BMT group or aGVHD group respectively. The colony-forming unit-fibroblast（CFU-F) experiment was performed to evaluate the potential effects of serums on the self-renewal ability of MSC. The expression of CD29 and CD105 of MSC was evaluated by immunofluorescence staining. In addition, the expression of self-renewal-related genes including Oct-4, Sox-2, and Nanog in MSC was detected by real-time fluorescence quantitative PCR（RT-qPCR). We successfully established an in vitro cell model that could mimic the bone marrow microenvironment damage of the mouse with aGVHD. CFU-F assay showed that, on day 7 after the culture, compared with the BMT group, MSC colony formation ability of aGVHD serum concentrations groups of 2% and 5% was significantly reduced （P < 0．05）; after the culture, at day 14, compared with the BMT group, MSC colony formation ability in different aGVHD serum concentration was significantly reduced （P < 0．05）. The immunofluorescence staining showed that, compared with the BMT group, the proportion of MSC surface molecules CD29+ and CD105+ cells was significantly dereased in the aGVHD serum concentration group (P < 0.05), the most significant difference was at a serum concentration of 10% （P < 0．001, P < 0.01）. The results of RT-qPCR detection showed that the expression of the MSC self-renewal-related genes Oct-4, Sox-2, and Nanog was decreased, the most significant difference was observed at an aGVHD serum concentration of 10% （P < 0.01,P < 0．001,P < 0．001）. By co-culturing different concentrations of mouse aGVHD serum and mouse MSC, we found that the addition of mouse aGVHD serum at different concentrations impaired the MSC self-renewal ability, which providing a new tool for the field of aGVHD bone marrow microenvironment damage.

MicroRNA-18a prevents senescence of mesenchymal stem cells by targeting CTDSPL.

Stem cell therapy requires massive-scale homogeneous stem cells under strict qualification control. However, Prolonged ex vivo expansion impairs the biological functions and results in senescence of mesenchymal stem cells (MSCs). We investigated the function of CTDSPL in the premature senescence process of MSCs and clarified that miR-18a-5p played a prominent role in preventing senescence of long-term cultured MSCs and promoting the self-renewal ability of MSCs. Over-expression of CTDSPL resulted in an enlarged morphology, up-regulation of p16 and accumulation of SA-β-gal of MSCs. The reduced phosphorylated RB suggested cell cycle arrest of MSCs. All these results implied that CTDSPL induced premature senescence of MSCs. We further demonstrated that miR-18a-5p was a putative regulator of CTDSPL by luciferase reporter assay. Inhibition of miR-18a-5p promoted the expression of CTDSPL and induced premature senescence of MSCs. Continuous overexpression of miR-18a-5p improved self-renewal of MSCs by reducing ROS level, increased expression of Oct4 and Nanog, and promoted growth rate and differentiation capability. We reported for the first time that the dynamic interaction of miR-18a-5p and CTDSPL is crucial for stem cell senescence.

Mechanism of Bazi Bushen capsule in delaying the senescence of mesenchymal stem cells based on network pharmacology and experimental validation

Ageing is becoming an increasingly serious problem; therefore, there is an urgent need to find safe and effective anti-ageing drugs. AimsTo investigate the effects of Bazi Bushen capsule (BZBS) on the senescence of mesenchymal stem cells (MSCs) and explore its mechanism of action. MethodsNetwork pharmacology was used to predict the targets of BZBS in delaying senescence in MSCs. For in vitro studies, MSCs were treated with D-gal, BZBS, and NMN, and cell viability, cell senescence, stemness-related genes, and cell cycle were studied using cell counting kit-8 (CCK-8) assay, SA-β-galactosidase (SA-β-gal) staining, Quantitative Real-Time PCR (qPCR) and flow cytometry (FCM), respectively. Alkaline phosphatase (ALP), alizarin red, and oil red staining were used to determine the osteogenic and lipid differentiation abilities of MSCs. Finally, the expression of senescence-related genes and cyclin-related factors was detected by qPCR and western blotting. ResultsNetwork pharmacological analysis suggested that BZBS delayed cell senescence by interfering in the cell cycle. Our in vitro studies suggested that BZBS could significantly increase cell viability (P < 0.01), decrease the quantity of β-galactosidase+ cells (P < 0.01), downregulate p16 and p21 (P < 0.05, P < 0.01), improve adipogenic and osteogenic differentiation, and upregulate Nanog, OCT4 and SOX2 genes (P < 0.05, P < 0.01) in senescent MSCs. Moreover, BZBS significantly reduced the proportion of senescent MSCs in the G0/G1 phase (P < 0.01) and enhanced the expression of CDK4, Cyclin D1, and E2F1 (P < 0.05, P < 0.01, respectively). Upon treatment with HY-50767A, a CDK4 inhibitor, the upregulation of E2F1 was no longer observed in the BZBS group. ConclusionsBZBS can protect MSCs against D-gal-induced senescence, which may be associated with cell cycle regulation via the Cyclin D1/CDK4/E2F1 signalling pathway.

Platelet-rich plasma-derived exosomes boost mesenchymal stem cells to promote peripheral nerve regeneration

Peripheral nerve injury (PNI) remains a severe clinical problem with debilitating consequences. Mesenchymal stem cell (MSC)-based therapy is promising, but the problems of poor engraftment and insufficient neurotrophic effects need to be overcome. Herein, we isolated platelet-rich plasma-derived exosomes (PRP-Exos), which contain abundant bioactive molecules, and investigated their potential to increase the regenerative capacity of MSCs. We observed that PRP-Exos significantly increased MSC proliferation, viability, and mobility, decreased MSC apoptosis under stress, maintained MSC stemness, and attenuated MSC senescence. In vivo, PRP-Exo-treated MSCs (pExo-MSCs) exhibited an increased retention rate and heightened therapeutic efficacy, as indicated by increased axonal regeneration, remyelination, and recovery of neurological function in a PNI model. In vitro, pExo-MSCs coculture promoted Schwann cell proliferation and dorsal root ganglion axon growth. Moreover, the increased neurotrophic behaviour of pExo-MSCs was mediated by trophic factors, particularly glia-derived neurotrophic factor (GDNF), and PRP-Exos activated the PI3K/Akt signalling pathway in MSCs, leading to the observed phenotypes. These findings demonstrate that PRP-Exos may be novel agents for increasing the ability of MSCs to promote neural repair and regeneration in patients with PNI.

Shilajit Boosts Osteogenic Ability of Mesenchymal Stem Cells for Regeneration of Rat Bone Defect

Shilajit Boosts Osteogenic Ability of Mesenchymal Stem Cells for Regeneration of Rat Bone Defect

Exploring the Impact of Leishmania Major on Mesenchymal Stem Cells: Evaluating Differentiation, and Immunomodulatory Function.

Pathogen recognition receptors (PRRs), which play a crucial role in responding to pathogens, affect the function of mesenchymal stem cells (MSCs). One important group of PRRs is the toll-like receptors (TLRs). When PRRs are activated, they can alter the expression of specific surface markers, the ability of MSCs to differentiate, and the types of substances they secrete. These modifications in MSC function may have unexpected consequences for patients. In this study, we examined how Leishmania major (L. major) promastigotes affect the properties of MSCs. MSCs were isolated from adipose tissue and categorized into two groups: one group left untreated and the other group exposed to L. major. Giemsa staining was employed to accurately quantify the number of parasites that entered the cells. After 72 hours, real-time polymerase chain reaction was utilized to assess the expression of TLRs. Additionally, the flow cytometry technique was used to evaluate the expression of surface markers on the MSCs. Our results showed that MSCs can engulf parasites and increase the expression of TLR4 and TLR6. The pro-inflammatory cytokine increased, and the transforming growth factor-β decreased significantly. The parasite exposure increased reactive oxygen species production. Additionally, the percentage of cluster differentiation (CD) 73 decreased, and the mean fluorescent index of CD29 and CD73 was down-regulated by L. major. Exposure to parasites diminishes the immunomodulatory capacity of MSCs. This discovery holds significance for the application of MSCs in addressing parasite infections and underscores the need for additional research to enhance their therapeutic effectiveness.

Rebuilding the myocardial microenvironment to enhance mesenchymal stem cells-mediated regeneration in ischemic heart disease.

Mesenchymal stem cells (MSCs) are naturally-derived regenerative materials that exhibit significant potential in regenerative medicine. Previous studies have demonstrated that MSCs-based therapy can improve heart function in ischemia-injured hearts, offering an exciting therapeutic intervention for myocardial ischemic infarction, a leading cause of worldwide mortality and disability. However, the efficacy of MSCs-based therapies is significantly disturbed by the myocardial microenvironment, which undergoes substantial changes following ischemic injury. After the ischemic injury, blood vessels become obstructed and damaged, and cardiomyocytes experience ischemic conditions. This activates the hypoxia-induced factor 1 (HIF-1) pathway, leading to the rapid production of several cytokines and chemokines, including vascular endothelial growth factor (VEGF) and stromal-derived factor 1 (SDF-1), which are crucial for angiogenesis, cell migration, and tissue repair, but it is not sustainable. MSCs respond to these cytokines and chemokines by homing to the injured site and participating in myocardial regeneration. However, the deteriorated microenvironment in the injured myocardium poses challenges for cell survival, interacting with MSCs, and constraining their homing, retention, and migration capabilities, thereby limiting their regenerative potential. This review discusses how the deteriorated microenvironment negatively affects the ability of MSCs to promote myocardial regeneration. Recent studies have shown that optimizing the microenvironment through the promotion of angiogenesis can significantly enhance the efficacy of MSCs in treating myocardial infarction. This approach harnesses the full therapeutic potential of MSCs-based therapies for ischemic heart disease.

IFN-γ-Licensed Mesenchymal Stem Cells Are More Susceptible to Death when Exposed to Quorum-Sensing Signal Molecule OdDHL and Less Effective in Inhibiting the Growth of Pseudomonas aeruginosa.

Currently, a series of licensing strategies has been investigated to enhance the functional properties of mesenchymal stem cells (MSCs). Licensing with IFN-γ is one of the most investigated strategies for enhancing the immunosuppressive potential of such cells. However, it is not yet known whether this licensing strategy could interfere with the ability of MSCs to control bacterial growth, which may be relevant considering their clinical potential. In this study, we compared the antimicrobial potential of IFN-γ-licensed and unlicensed MSCs by exposing them to Pseudomonas aeruginosa and its quorum-sensing inducer molecule OdDHL. Our data show that-when challenged with OdDHL-IFN-γ-licensed and unlicensed MSCs present increased levels of the antimicrobial HAMP transcript, but that only IFN-γ-licensed MSCs undergo modulation of CASP1 and BCL2, entering apoptosis. Furthermore, we demonstrate that only IFN-γ-licensed MSCs show modulation in genes involved in apoptosis and tend to undergo cell death when cultured with P. aeruginosa. As a consequence, IFN-γ-licensed MSCs showed lower capacity to control bacterial growth, compared to unlicensed MSCs. Taken together, our observations reveal an increased susceptibility to apoptosis of IFN-γ-licensed MSCs, which compromises their potential to control the bacterial growth in vitro. These findings are relevant to the field of cell therapy, considering the potential applicability of MSCs.