Given the high heterogeneity of type 2 diabetes mellitus (T2DM), it is imperative to develop personalized stem cell infusion regimen for targeted metabolic phenotype in order to ensure optimal therapeutic efficacy. In this study, we conducted a comparative analysis of 4 infusion regimens involving single and repeated infusions of human umbilical cord Wharton's jelly-derived MSCs (hucMSCs), single infusions of umbilical cord blood mononuclear cells (UCB), and sequential infusions of hucMSCs and UCB in T2DM rats. Results showed all 4 infusion regimens exhibited comparable efficacy in lowering fasting blood glucose levels and suppressing glucagon secretion. Single and double infusions of hucMSCs exhibited a tendency to migrate to the liver, thereby better at ameliorating hepatic glucose metabolism by enhancing glycogen synthesis and storage, promoting glycolysis, inhibiting gluconeogenesis, and improving insulin signal transduction. The sequential infusion of hucMSCs and UCB demonstrated specific cell tropism toward the pancreas, leading to prolonged glucose-lowering effects following a glucose tolerance test, restoration of early-phase insulin secretion, stimulation of islet beta cell proliferation and improvement in the beta/alpha ratio. Multiple injections, regardless of cell type, reduced the expression of systemic chronic inflammatory markers such as IL-1β, IL-6, IL-17, IL-22, and IFN-γ. Finally, a single dose of UCB exhibited a greater tendency to target visceral fat and enhanced effectiveness in regulating levels of total cholesterol and triglycerides. In conclusion, our study provided personalized stem cell regimens for diverse T2DM metabolic phenotypes, thereby offering improved treatment alternatives for future clinical trials and applications.

Mesenchymal stem cells (MSCs) have shown potential in treating immune-mediated diseases due to their immunomodulatory properties, which can be enhanced by priming with inflammatory cytokines like interferon-gamma (IFN-γ). This study evaluates the therapeutic effects of IFN-γ-primed canine adipose tissue-derived MSCs (AMSCs) in a mouse model of inflammatory bowel disease (IBD). Canine AMSCs were primed with 50 ng/mL recombinant canine IFN-γ for 48 h, and the effects were compared to those seen in naïve (unprimed) AMSCs. IBD was induced in mice using dextran sodium sulfate (DSS), and AMSCs were injected intraperitoneally on days 1 and 3. The mice treated with IFN-γ-primed AMSCs showed improved clinical outcomes, including a reduced disease activity index (DAI), less body weight loss, and longer colon length compared to the mice treated with naïve AMSCs. A histological analysis revealed less damage to the intestinal structures and reduced inflammatory cell infiltration. IFN-γ priming led to a shift in the immune cell balance in the gut, decreasing pro-inflammatory macrophages (Ly6Chi) and increasing anti-inflammatory macrophages (Ly6Clo/MHC-IIhi). This was associated with the reduced expression of inflammatory cytokine genes (Il-1β, Il-6, and Il-18) and increased expression of the intestinal stem cell marker Lgr5. These findings suggest that IFN-γ-primed AMSCs offer enhanced therapeutic potential for treating CE in veterinary medicine.

The present study was aimed to explore the possible mechanisms by which caprine Wharton's jelly-derived MSCs (WJ-MSCs) perform their immunomodulatory function. WJ-MSCs were isolated through explants culture and characterized as per ISCT criteria using culture behavior, expression of surface markers by PCR, FACS and immunocytochemical localization (ICC), trilineage differentiation potential etc. Secretory behavior for important biomolecules (IDO, TGFβ1, VEGF, IL6) was evaluated by ICC and western blot assay. Cell-to-cell communication was studied by culturing cells in cell-cell contact and trans-well system. The MSCs when co-cultured with activated Tc and Th cells, down-regulation of T cell cytokine as well as upregulation of immunomodulatory factors (VEGF A, IL10, IL6, IDO, iNOS, PTGS2, HGF, TGFβ, CXCL10, CXCL11) was noticed in both cell-cell contact and trans-well culture system which was significantly higher in cell-cell contact system. Trilineage differentiation of MSCs showed significant upregulation of MHC I (CAHI) and MHC II (CLA DRB3) molecules suggesting better clinical applications of MSCs without differentiation to avoid immune rejection. It can be concluded that WJ-MSCs perform their immunomodulation through the secretion of a battery of biomolecules and work in both cell-cell contact manner and through their secretome.

Alzheimer's disease (AD) develops as a result of oxidative damage to neurons and chronic inflammation of microglia. These processes can be influenced by the use of a conditioned medium (CM) derived from mesenchymal stem cells (MSCs). The CM contains a wide range of factors that have neurotrophic, antioxidant, and anti-inflammatory effects. In addition, the therapeutic potential of the CM can be further enhanced by pretreating the MSCs to increase their paracrine activity. The current study aimed to investigate the neuroprotective effects of CM derived from MSCs, which were either activated by a TLR3 ligand or exposed to CoCl2, a hypoxia mimetic (pCM or hCM, respectively), in an in vitro model of AD.

We have developed a novel in vitro model of AD that allows us to investigate the neuroprotective and anti-inflammatory effects of MSCs on induced neurodegeneration in the PC12 cell line and the activation of microglia using THP-1 cells.

This study demonstrates for the first time that pCM and hCM exhibit more pronounced immunosuppressive effects on proinflammatory M1 macrophages compared to CM derived from untreated MSCs (cCM). This may help prevent the development of neuroinflammation by balancing the M1 and M2 microglial phenotypes via the decreased secretion of proinflammatory cytokines (IL-1β, IL-6, and TNF-α) and increased secretion of IL-4, as well as the expression of IL-10 and TGF-β by macrophages. Moreover, a previously unknown increase in the neurotrophic properties of hCM was discovered, which led to an increase in the viability of neuron-like PC12 cells under H2O2-induced oxidative-stress conditions. These results are likely associated with an increase in the production of growth factors, including vascular endothelial growth factor (VEGF). In addition, the neuroprotective effects of CM from preconditioned MSCs are also mediated by the activation of the Nrf2/ARE pathway in PC12 cells.

TLR3 activation in MSCs leads to more potent immunosuppressive effects of the CM against pro-inflammatory M1 macrophages, while the use of hCM led to increased neurotrophic effects after H2O2-induced damage to neuronal cells. These results are of interest for the potential treatment of AD with CM from preactivated MSCs.

Mesenchymal stem cells (MSCs) are a promising tool that may be used in regenerative medicine. Thanks to their ability to differentiate and paracrine signaling, they can be used in the treatment of many diseases. Undifferentiated MSCs can support the regeneration of surrounding tissues through secreted substances and exosomes. This is possible thanks to the production of growth factors. These factors stimulate the growth of neighboring cells, have an anti-apoptotic effect, and support angiogenesis, and MSCs also have an immunomodulatory effect. The level of secreted factors may vary depending on many factors. Apart from the donor's health condition, it is also influenced by the source of MSCs, methods of harvesting, and even the banking of cells. This work is a review of research on how the patient's health condition affects the properties of obtained MSCs. The review discusses the impact of the patient's diabetes, obesity, autoimmune diseases, and inflammation, as well as the impact of the source of MSCs and methods of harvesting and banking cells on the phenotype, differentiation capacity, anti-inflammatory, angiogenic effects, and proliferation potential of MSCs. Knowledge about specific clinical factors allows for better use of the potential of stem cells and more appropriate targeting of procedures for collecting, multiplying, and banking these cells, as well as for their subsequent use. This article aims to review the characteristics, harvesting, banking, and paracrine signaling of MSCs and their role in diabetes, obesity, autoimmune and inflammatory diseases, and potential role in regenerative medicine.

Finding novel treatments for cardiovascular diseases (CVDs) is a hot topic in medicine; cell-based therapies have reported promising news for controlling dangerous complications of heart disease such as myocardial infarction (MI) and heart failure (HF). Various progenitor/stem cells were tested in various in-vivo, in-vitro, and clinical studies for regeneration or repairing the injured tissue in the myocardial to accelerate the healing. Fetal, adult, embryonic, and induced pluripotent stem cells (iPSC) have revealed the proper potency for cardiac tissue repair. As an essential communicator among cells, exosomes with specific contacts (proteins, lncRNAs, and miRNAs) greatly promote cardiac rehabilitation. Interestingly, stem cell-derived exosomes have more efficiency than stem cell transplantation. Therefore, stem cells induced pluripotent stem cells (iPSCs), embryonic stem cells (ESCs), cardiac stem cells (CDC), and skeletal myoblasts) and their-derived exosomes will probably be considered an alternative therapy for CVDs remedy. In addition, stem cell-derived exosomes have been used in the diagnosis/prognosis of heart diseases. In this review, we explained the advances of stem cells/exosome-based treatment, their beneficial effects, and underlying mechanisms, which will present new insights in the clinical field in the future.

The therapeutic efficacy of mesenchymal stromal cells (MSCs) in inflammatory bowel disease is not completely known and is not consistent. Priming with inflammatory cytokines has been proposed to adapt MSCs to an inflammatory environment to have them ready to counteract it, but may have undesirable effects on MSCs, such as increased immunogenicity. In this study, we hypothesized that priming MSCs with inflamed intestinal tissue would more effectively enhance their therapeutic effect on intestinal inflammation.

The capacity of canine adipose-derived MSCs (cADSCs) primed with colon tissue homogenates from mice with experimentally induced colitis or a combination of tumor necrosis factor-α and interferon-γ to inhibit T-cell proliferation was analyzed, along with their own apoptosis, proliferation, cell surface marker expression, and transcriptome. In addition, colitis mice were treated with the primed cADSCs to assess colitis severity and immune cell profile.

Priming with cytokines induced apoptosis, decreased cell proliferation, and major histocompatibility complex-II gene expression in cADSCs, but these adverse effects were mild or absent with colitis-tissue priming. cADSCs primed with colitis tissue reduced the severity of colitis via the induction of M2 macrophages and T-regulatory cells and suppression of T-helper (Th)1/Th17-cell responses, and their effects were comparable to those of cytokine-primed cells.

Our results emphasize the importance of the activation of MSCs by the appropriate microenvironment to maximize their therapeutic effect.

Wharton's jelly mesenchymal stem cells (WJ-MSCs) are gaining significant attention in regenerative medicine for their potential to treat degenerative diseases and mitigate radiation injuries. WJ-MSCs are more naïve and have a better safety profile, making them suitable for both autologous and allogeneic transplantations. This review highlights the regenerative potential of WJ-MSCs and their clinical applications in mitigating various types of radiation injuries. In this review, we will also describe why WJ-MSCs will become one of the most probable stem cells for future regenerative medicine along with a balanced view on their strengths and weaknesses. Finally, the most updated literature related to both preclinical and clinical usage of WJ-MSCs for their potential application in the regeneration of tissues and organs will also be compiled.

Multipotent mesenchymal stromal cells (MSCs) hold promises for cell therapy and tissue engineering due to their self-renewal and differentiation abilities, along with immunomodulatory properties and trophic factor secretion. Extracellular vesicles (EVs) from MSCs offer similar therapeutic effects. However, MSCs are heterogeneous and lead to variable outcomes. In vitro priming enhances MSC performance, improving immunomodulation, angiogenesis, proliferation, and tissue regeneration. Various stimuli, such as cytokines, growth factors, and oxygen tension, can prime MSCs. Two classical priming methods, interferon-gamma (IFN-γ) and hypoxia, enhance MSC immunomodulation, although standardized protocols are lacking. This review discusses priming protocols, highlighting the most commonly used concentrations and durations, along with mechanisms and in vivo therapeutics effects of primed MSCs and their EVs. The feasibility of up-scaling their production was also discussed. The review concluded that priming with IFN-γ or hypoxia (alone or in combination with other factors) boosted the immunomodulation capability of MSCs and their EVs, primarily via the JAK/STAT and PI3K/AKT and Leptin/JAK/STAT and TGF-β/Smad signalling pathways, respectively. Incorporating priming in MSC and EV production enables translation into cell-based or cell-free therapies for various disorders.

Human umbilical cord mesenchymal stem cell (hucMSC)-derived exosomes (Exo) have been frequently investigated for disease control. This study was designed to explore the effects of hucMSC-Exo carrying lncRNA family with sequence similarity 99-member B (Exo-lncRNA FAM99B) on hepatocellular carcinoma (HCC) cell behaviour. The expression of lncRNA FAM99B in HCC cells was measured by reverse-transcription quantitative polymerase chain reaction. Protein levels of exosomal markers were quantified using western blotting. Flow cytometry analyses were performed to detect surface markers of hucMSCs and to measure the effects of Exo-lncRNA FAM99B on HCC cell cycle progression and cell apoptosis. Nanoparticle tracking analysis was used to measure the particle size of the exosomes. Additionally, cell viability was evaluated using methyl thiazolyl tetrazolium assays, and Transwell assays were performed to measure cell migration and invasion. Xenograft tumor models were established to explore the role of Exo-lncRNA FAM99B in vivo. Experimental results revealed that lncRNA FAM99B was downregulated in HCC cell lines, and low level of FAM99B is associated with poor survival rates in patients with HCC according to bioinformatics analysis. HucMSCs were identified in a good morphology with positively expressed CD105, CD29, and CD44 as well as negatively expressed CD31, CD14, and HLA-DR. High protein levels of exosomal markers (Alix, CD63 and TSG101) identified the existence of HucMSC-Exo. Importantly, the hucMSCs-Exo could enter HCC cells and exerted a suppressive effect on malignant cell activities. Moreover, overexpression of Exo-lncRNA FAM99B enhanced cell cycle arrest and cell apoptosis while suppressing cell viability, migration, and invasion in HCC. Exo-siRNA-FAM99B exerted the opposite effects on HCC cell process. In vivo experiments verified that Exo-lncRNA FAM99B inhibited tumorigenesis in HCC. In summary, lncRNA FAM99B derived from hucMSC-Exo inhibited malignant cellular phenotypes and tumorigenesis in HCC, which might provide a novel therapeutic strategy for HCC treatment.

Hematopoiesis is a process by which all blood cells are formed. The mechanisms controlling it have been studied for decades. Surprisingly, while hematopoietic stem cells are among the most extensively studied stem cell types, the complete understanding of how they are regulated during development, adulthood, or in non-homeostatic conditions remains elusive. In this review, our primary focus is on research findings that explore where hematopoietic precursors are found in adults outside their primary niches in the bone marrow. This phenomenon is termed extramedullary hematopoiesis (EMH). Early in development hematopoietic stem cells migrate through different regions within and outside the embryo and later the fetus. Although, the primary home for hematopoietic progenitors is the adult bone marrow, it is now recognized that other adult organs may act as hematopoietic progenitor reservoirs both in mice and humans. The first reports about this topic were principally originated from clinical observations, in cases where the bone marrow was malfunctioning, leading to an aberrant hematopoiesis outside the bone marrow. It is worth highlighting that those extramedullary organs, like the small intestine or fat tissue, contain subsets of fully functioning hematopoietic progenitors demonstrated by both in vitro and in vivo studies. Nonetheless, there are still some unanswered questions regarding the source of these cells, how they differ in function compared to their counterparts in the bone marrow, and the specific roles they play within the tissues where they are located.

The adnexa fetal tissues are sources of mesenchymal stromal cells (MSCs) due to their noninvasive harvest, with all biological material discarded most of the time. MSCs are a promise regarding to their plasticity, self-renewal, differentiation potentials, immunomodulatory and anti-inflammatory properties, which have made clinical stem cell therapy a reality. The present study aimed to characterize and evaluate the immunomodulation ability of bovine mesenchymal cells collected from bovine amniotic fluid (bAFMSCs) isolated and subjected to sixth consecutive culture passages in vitro. The multilineage properties of the bAFMSCs collections confirmed the ability to undergo adipogenic, chondrogenic and osteogenic differentiation. The mesenchymal gene transcription CD106, CD73, CD29, CD90 and CD166 were detected in bAFMSCs, whereas CD34 and CD45 were not detected. Regarding cytokine mRNA expression, IL2, IL6, INFα, INFβ, INFγ, TNFα and TNFβ were downregulated, while IL10 was highly regulated in all studied passages. The present study demonstrated the immunological properties and multipotency of in vitro bAFMSCs collections, and thus, they can be tested in cattle pathological treatments or multiplication by nuclear transfer cloning.

Hematopoiesis is a tightly regulated process that produces all adult blood cells and immune cells from multipotent hematopoietic stem cells (HSCs). HSCs usually remain quiescent, and in the presence of external stimuli like infection or inflammation, they undergo division and differentiation as a compensatory mechanism. Normal hematopoiesis is impacted by systemic inflammation, which causes HSCs to transition from quiescence to emergency myelopoiesis. At the molecular level, inflammatory cytokine signaling molecules such as tumor necrosis factor (TNF), interferons, interleukins, and toll-like receptors can all cause HSCs to multiply directly. These cytokines actively encourage HSC activation, proliferation, and differentiation during inflammation, which results in the generation and activation of immune cells required to combat acute injury. The bone marrow niche provides numerous soluble and stromal cell signals, which are essential for maintaining normal homeostasis and output of the bone marrow cells. Inflammatory signals also impact this bone marrow microenvironment called the HSC niche to regulate the inflammatory-induced hematopoiesis. Continuous pro-inflammatory cytokine and chemokine activation can have detrimental effects on the hematopoietic system, which can lead to cancer development, HSC depletion, and bone marrow failure. Reactive oxygen species (ROS), which damage DNA and ultimately lead to the transformation of HSCs into cancerous cells, are produced due to chronic inflammation. The biological elements of the HSC niche produce pro-inflammatory cytokines that cause clonal growth and the development of leukemic stem cells (LSCs) in hematological malignancies. The processes underlying how inflammation affects hematological malignancies are still not fully understood. In this review, we emphasize the effects of inflammation on normal hematopoiesis, the part it plays in the development and progression of hematological malignancies, and potential therapeutic applications for targeting these pathways for therapy in hematological malignancies.

A wound takes a long time to heal and involves several steps. Following tissue injury, inflammation is the primary cause of tissue regeneration and repair processes. As a result, the pathophysiological processes involving skin damage, healing, and remodeling depend critically on the control of inflammation. The fact that it is a feasible target for improving the prognosis of wound healing has lately become clear. Mesenchymal stem cells (MSCs) are an innovative and effective therapeutic option for wound healing due to their immunomodulatory and paracrine properties. By controlling the inflammatory milieu of wounds through immunomodulation, transplanted MSCs have been shown to speed up the healing process. In addition to other immunomodulatory mechanisms, including handling neutrophil activity and modifying macrophage polarization, there may be modifications to the activation of T cells, natural killer (NK) cells, and dendritic cells (DCs). Furthermore, several studies have shown that pretreating MSCs improves their ability to modulate immunity. In this review, we summarize the existing knowledge about how MSCs influence local inflammation in wounds by influencing immunity to facilitate the healing process. We also provide an overview of MSCs optimizing techniques when used to treat wounds.

Promising preclinical stroke research has not yielded meaningful and significant success in clinical trials. This lack of success has prompted the need for refinement of preclinical studies with the intent to optimize the chances of clinical success. Regenerative medicine, especially using mesenchymal stem/stromal cells (MSCs), has gained popularity in the last decade for treating many disorders, including central nervous system (CNS), such as stroke. In addition to less stringent ethical constraints, the ample availability of MSCs also makes them an attractive alternative to totipotent and other pluripotent stem cells. The ability of MSCs to differentiate into neurons and other brain parenchymal and immune cells makes them a promising therapy for stroke. However, these cells also have some drawbacks that, if not addressed, will render MSCs unfit for treating ischaemic stroke. In this review, we highlighted the molecular and cellular changes that occur following an ischaemic stroke (IS) incidence and discussed the physiological properties of MSCs suitable for tackling these changes. We also went further to discuss the major drawbacks of utilizing MSCs in IS and how adequate priming using both hypoxia and interleukin-1 can optimize the beneficial properties of MSCs while eliminating these drawbacks.

Acute radiation syndrome (ARS) is also known as triple syndrome, which develops after whole-body radiation exposure. During unforeseen exposures, these syndromes are set in depending on the dose of radiation. Cell-based therapy, especially using stem cells and their soluble factors, is gaining wide attention in the field of regenerative medicine to treat various diseases, including degenerative diseases. Stem cells attract prime attention because of their profound inherent tissue repair capability and regeneration potential. Further, stem cell therapy can be one of the promising strategies for the amelioration of ARS because of its ability to lodge in damaged tissue and release regenerative cytokines by sensing the local injury. In this regard, human Wharton's jelly-derived mesenchymal stem cells (hWJ-MSCs) have gained substantial attention for their applications in the treatment of various human diseases due to several advantages offered by them. This article is intended to provide future perspective on the use of WJ-MSCs for the management of accidental radiation injury in pre-clinical models, and finally, their utility in regeneration of damaged tissues and organs.

Repeated Ovum Pick Up (OPU) could have a detrimental effect on ovarian function, reducing In Vitro Embryo Production (IVEP). The present study examined the therapeutic effect of adipose-derived Mesenchymal Stem Cells (MSCs) or its Conditioned Medium (ConM) on ovarian trauma following repeated OPU. Resolvin E1 (RvE1) and Interleukin-12 (IL-12) were investigated as biomarkers.

Jersey heifers (n=8) experienced 11 OPU sessions including 5 pre-treatment and 6 treatment sessions. Heifers received intra-ovarian administration of MSCs or ConM (right ovary) and Dulbecco's Modified Phosphate Buffer Saline (DMPBS; left ovary) after OPU in sessions 5 and 8 and 2 weeks after session 11. The concentrations of RvE1 and IL-12 in follicular fluid was evaluated on sessions 1, 5, 6, 9, and 4 weeks after session 11. Following each OPU session, the IVEP parameters were recorded.

Intra-ovarian administration of MSCs, ConM, and DMPBS did not affect IVEP parameters (p>0.05). The concentration of IL-12 in follicular fluid increased at the last session of pre-treatment (Session 5; p<0.05) and remained elevated throughout the treatment period. There was no correlation between IL-12 and IVEP parameters (p>0.05). However, RvE1 remained relatively high during the pre-treatment and decreased toward the end of treatment period (p<0.05). This in turn was associated with decline in some IVEP parameters (p<0.05).

Intra-ovarian administration of MSCs or ConM during repeated OPU did not enhance IVEP outcomes in Bos taurus heifers. The positive association between RvE1 and some of IVEP parameters could nominate RvE1 as a promising biomarker to predict IVEP parameters following repeated OPU.

Mesenchymal stromal cells (MSCs) are polymorphic, adherent cells with the capability to stimulate tissue regeneration and modulate immunity. MSCs have been broadly investigated for potential therapeutic applications, particularly immunomodulatory properties, wound healing and tissue regeneration. The exact physiologic role of MSCs, however, remains poorly understood, and this gap in knowledge significantly impedes the rational development of therapeutic cells. Here, we considered interferon γ (IFN-γ) and tumor necrosis factor alpha (TNF-α), two cytokines likely encountered physiologically and commonly used in cell manufacturing. For comparison, we studied interleukin-10 (IL-10) (anti-inflammatory) and interleukin-4 (IL-4) (type 2 cytokine).

We directly assessed the effects of these cytokines on bone marrow MSCs by comparing RNA Seq transcriptional profiles. Western blotting and flow cytometry were also used to evaluate effects of cytokine priming.

The type 1 cytokines (IFN-γ and TNF-α) induced striking changes in gene expression and remarkably different profiles from one another. Importantly, priming MSCs with either of these cytokines did not increase variability among multiple donors beyond what is intrinsic to non-primed MSCs from different donors. IFN-γ-primed MSCs expressed IDO1 and chemokines that recruit activated T cells. In contrast, TNF-α-primed MSCs expressed genes in alternate pathways, namely PGE2 and matrix metalloproteinases synthesis, and chemokines that recruit neutrophils. IL-10 and IL-4 priming had little to no effect.

Our data suggest that IFN-γ-primed MSCs may be a more efficacious immunosuppressive therapy aimed at diseases that target T cells (ie, graft-versus-host disease) compared with TNF-α-primed or non-primed MSCs, which may be better suited for therapies in other disease settings. These results contribute to our understanding of MSC bioactivity and suggest rational ex vivo cytokine priming approaches for MSC manufacturing and therapeutic applications.

Mesenchymal stromal cells (MSCs) have regenerative and immunomodulatory properties, making them suitable for cell therapy. Toll-like receptors (TLRs) in MSCs respond to viral load by secreting immunosuppressive or proinflammatory molecules. The expression of anti-inflammatory molecules in MSCs can be altered by the concentration and duration of exposure to the TLR3 ligand polyinosinic-polycytidylic acid (poly(I:C)). This study aimed to optimize the preconditioning of MSCs with poly(I:C) to increase immunosuppressive effects and to identify MSCs with activated TLR3 (prMSCs).

Flow cytometry and histochemical staining were used to analyze MSCs for immunophenotype and differentiation potential. MSCs were exposed to poly(I:C) at 1 and 10 μg/mL for 1, 3, and 24 h, followed by determination of the expression of IDO1, WARS1, PD-L1, TSG-6, and PTGES2 and PGE2 secretion. MSCs and prMSCs were cocultured with intact (J-) and activated (J+) Jurkat T cells. The proportion of proliferating and apoptotic J+ and J- cells, IL-10 secretion, and IL-2 production after cocultivation with MSCs and prMSCs were measured. Liquid chromatography-mass spectrometry and bioinformatics analysis identified proteins linked to TLR3 activation in MSCs.

Poly(I:C) at 10 μg/mL during a 3-h incubation caused the highest expression of immunosuppression markers in MSCs. Activation of prMSCs caused a 18% decrease in proliferation and a one-third increase in apoptotic J+ cells compared to intact MSCs. Cocultures of prMSCs and Jurkat cells had increased IL-10 and decreased IL-2 in the conditioned medium. A proteomic study of MSCs and prMSCs identified 53 proteins with altered expression. Filtering the dataset with Gene Ontology and Reactome Pathway revealed that poly(I:C)-induced proteins activate the antiviral response. Protein‒protein interactions by String in prMSCs revealed that the antiviral response and IFN I signaling circuits were more active than in native MSCs. prMSCs expressed more cell adhesion proteins (ICAM-I and Galectin-3), PARP14, PSMB8, USP18, and GBP4, which may explain their anti-inflammatory effects on Jurkat cells.

TLR3 activation in MSCs is dependent on exposure time and poly(I:C) concentration. The maximum expression of immunosuppressive molecules was observed with 10 µg/mL poly(I:C) for 3-h preconditioning. This priming protocol for MSCs enhances the immunosuppressive effects of prMSCs on T cells.

Although it has been suggested that toll-like receptor (TLR) 3 and TLR4 activation alters mesenchymal stromal cells (MSCs)' immunoregulatory function as anti- or pro-inflammatory phenotypes, we have previously confirmed that TLR4-primed hUCB-MSCs alleviate lung inflammation and tissue injury in an E. coli-induced acute lung injury (ALI) mouse model. Therefore, we hypothesized that strong stimulation of TLR3 or TLR4 prompts hUCB-MSCs to exhibit an anti-inflammatory phenotype mediated by extracellular vesicles (EVs). In this study, we compared the anti-inflammatory effect of TLR3-primed and TLR4-primed hUCB-MSCs against an LPS-induced ALI in vitro model by treating MSCs, MSC-derived conditioned medium (CM), and MSC-derived extracellular vesicles (EVs). LPS-induced rat primary alveolar macrophage and RAW 264.7 cells were treated with naïve, TLR3-, and TLR4-primed MSCs and their derived CM and EVs. Flow cytometry and ELISA were used to evaluate M1-M2 polarization of macrophages and pro-inflammatory cytokine levels, respectively. LPS-stimulated macrophages showed significantly increased pro-inflammatory cytokines compared to those of the normal control, and the percentage of M2 macrophage phenotype was predominantly low. In reducing the inflammatory cytokines and enhancing M2 polarization, TLR3- and TLR4-primed MSCs were significantly more effective than the naïve MSCs, and this finding was also observed with the treatment of MSC-derived CMs and EVs. No significant difference between the efficacy of TLR3- and TLR-primed MSCs was observed. Strong stimulation of TLR3- and TLR4-stimulated hUCB-MSCs significantly reduced pro-inflammatory cytokine secretion from LPS-induced macrophages and significantly enhanced the M2 polarization of macrophages. We further confirmed that TLR-primed MSC-derived EVs can exert anti-inflammatory and immunosuppressive effects alone comparable to MSC treatment. We hereby suggest that in the LPS-induced macrophage in vitro model, EVs derived from both TLR3 and TLR4-primed MSCs can be a therapeutic candidate by promoting the M2 phenotype.

Mesenchymal stem/stromal cells (MSCs), a class of cells with proliferative, immunomodulatory, and reparative functions, have shown therapeutic potential in a variety of systemic diseases, including metabolic syndrome (MetS). The cluster of morbidities that constitute MetS might be particularly amenable for the application of MSCs, which employ an arsenal of reparative actions to target multiple pathogenic pathways simultaneously. Preclinical studies have shown that MSCs can reverse pathological changes in MetS mainly by inhibiting inflammation, improving insulin resistance, regulating glycolipid metabolism, and protecting organ function. However, several challenges remain to overcome before MSCs can be applied for treating MetS. For example, the merits of autologous versus allogeneic MSCs sources remain unclear, particularly with autologous MSCs obtained from the noxious MetS milieu. The distinct characteristics and relative efficacy of MSCs harvested from different tissue sources also require clarification. Moreover, to improve the therapeutic efficacy of MSCs, investigators have explored several approaches that improved therapeutic efficacy but may involve potential safety concerns. This review summarized the potentially useful MSCs strategy for treating MetS, as well as some hurdles that remain to be overcome. In particular, larger-scale studies are needed to determine the therapeutic efficacy and safety of MSCs for clinical application.

Mesenchymal stromal cells (MSCs) have broad potential as a cell therapy including for the treatment of drug-resistant inflammatory conditions with abnormal T cell proliferation such as graft-versus-host disease (GVHD). Clinical success, however, has been complicated by the heterogeneity of culture-expanded MSCs as well as donor variability. Here, we devise culture conditions that promote expansion of MSCs with enhanced immunomodulatory functions both in vitro and in animal models of GVHD.

Human bone marrow-derived MSCs were expanded at high-confluency (MSCHC) and low-confluency state (MSCLC). Their immunomodulatory properties were evaluated with in vitro co-culture assays based on suppression of activated T cell proliferation and secretion of pro-inflammatory cytokines from activated T cells. Metabolic state of these cells was determined, while RNA sequencing was performed to explore transcriptome of these MSCs. Ex vivo expanded MSCHC or MSCLC was injected into human peripheral blood mononuclear cells (PBMC)-induced GVHD mouse model to determine their in vivo therapeutic efficacy based on clinical grade scoring, human CD45+ blood count and histopathological examination.

As compared to MSCLC, MSCHC significantly reduced both the proliferation of anti-CD3/CD28-activated T cells and secretion of pro-inflammatory cytokines upon MSCHC co-culture across several donors even in the absence of cytokine priming. Mechanistically, metabolic analysis of MSCHC prior to co-culture with activated T cells showed increased glycolytic metabolism and lactate secretion compared to MSCLC, consistent with their ability to inhibit T cell proliferation. Transcriptome analysis further revealed differential expression of immunomodulatory genes including TRIM29, BPIFB4, MMP3 and SPP1 in MSCHC as well as enriched pathways including cytokine-cytokine receptor interactions, cell adhesion and PI3K-AKT signalling. Lastly, we demonstrate in a human PBMC-induced GVHD mouse model that delivery of MSCHC showed greater suppression of inflammation and improved outcomes compared to MSCLC and saline controls.

Our study provides evidence that ex vivo expansion of MSCs at high confluency alters the metabolic and transcriptomic states of these cells. Importantly, this approach maximizes the production of MSCs with enhanced immunomodulatory functions without priming, thus providing a non-invasive and generalizable strategy for improving the use of MSCs for the treatment of inflammatory diseases.

Sepsis is caused by dysregulated immune response to severe infection and hyper inflammation plays a central role in worsening the disease. The immunomodulatory properties of mesenchymal stem cells (MSCs) have been evaluated as a therapeutic candidate for sepsis. Reconditioned monocytes (RM), generated from healthy human peripheral blood mononuclear cells (PBMCs) exhibit both macrophage and MSCs-like properties. RM were administered at different stages of sepsis in a mouse model. It reduced serum levels of IL6, MCP-1, IL-10, improved hypothermia, increased survival, and recovery from 0 to 66% when combined with antibiotics in the mouse model. The reduced human leucocyte antigen DR molecules expression on RM enables their co-culture with PBMCs of sepsis patients which resulted in reduced ROS production, and up-regulated TGF-β while down-regulating IL6, IL8, and IL-10 in-vitro. RM are potentially immunomodulatory, enhance survival in sepsis mouse model and modulate inflammatory behaviour of sepsis patient's PBMCs.

Mesenchymal stem cells (MSCs) are an attractive therapeutic tool for tissue engineering and regenerative medicine owing to their regenerative and trophic properties. The best-known and most widely used are bone marrow MSCs, which are currently being harvested and developed from a wide range of adult and perinatal tissues. MSCs from different sources are believed to have different secretion potentials and production, which may influence their therapeutic effects. To confirm this, we performed a quantitative proteomic analysis based on the TMT technique of MSCs from three different sources: Wharton's jelly (WJ), dental pulp (DP), and bone marrow (BM). Our analysis focused on MSC biological properties of interest for tissue engineering. We identified a total of 611 differentially expressed human proteins. WJ-MSCs showed the greatest variation compared with the other sources. WJ produced more extracellular matrix (ECM) proteins and ECM-affiliated proteins and proteins related to the inflammatory and immune response processes. BM-MSCs expressed more proteins involved in osteogenic, adipogenic, neuronal, or muscular differentiation and proteins involved in paracrine communication. Compared to the other sources, DP-MSCs overexpressed proteins involved in the exocytosis process. The results obtained confirm the existence of differences between WJ, DP, and BM-MSCs and the need to select the MSC origin according to the therapeutic objective sought.

Wound healing has long been a complex problem, especially in chronic wounds. Although debridement, skin grafting, and antimicrobial dressings have been used to treat chronic wounds, their treatment period is long, expensive, and has specific rejection reactions. The poor treatment results of traditional methods have caused psychological stress to patients and a substantial economic burden to society. Extracellular vesicles (EVs) are nanoscale vesicles secreted by cells. They play an essential role in intercellular communication. Numerous studies have confirmed that stem cell-derived extracellular vesicles (SC-EVs) can inhibit overactive inflammation, induce angiogenesis, promote re-epithelization, and reduce scar formation. Therefore, SC-EVs are expected to be a novel cell-free strategy for chronic wound treatment. We first summarize the pathological factors that hinder wound healing and discuss how SC-EVs accelerate chronic wound repair. And then, we also compare the advantages and disadvantages of different SC-EVs for chronic wound treatment. Finally, we discuss the limitations of SC-EVs usage and provide new thoughts for future SC-EVs research in chronic wound treatment.

Medicinal signaling cells (MSC) exhibit distinct molecular signatures and biological abilities, depending on the type of tissue they originate from. Recently, we isolated and described a new population of stem cells residing in the coronary corium, equine hoof progenitor cells (HPCs), which could be a new promising cell pool for the treatment of laminitis. Therefore, this study aimed to compare native populations of HPCs to well-established adipose-derived stem cells (ASCs) in standard culture conditions and in a pro-inflammatory milieu to mimic a laminitis condition. ASCs and HPCs were either cultured in standard conditions or subjected to priming with a cytokines cocktail mixture. The cells were harvested and analyzed for expression of key markers for phenotype, mitochondrial metabolism, oxidative stress, apoptosis, and immunomodulation using RT-qPCR. The morphology and migration were assessed based on fluorescent staining. Microcapillary cytometry analyses were performed to assess the distribution in the cell cycle, mitochondrial membrane potential, and oxidative stress. Native HPCs exhibited a similar morphology to ASCs, but a different phenotype. The HPCs possessed lower migration capacity and distinct distribution across cell cycle phases. Native HPCs were characterized by different mitochondrial dynamics and oxidative stress levels. Under standard culture conditions, HPCs displayed different expression patterns of apoptotic and immunomodulatory markers than ASCs, as well as distinct miRNA expression. Interestingly, after priming with the cytokines cocktail mixture, HPCs exhibited different mitochondrial dynamics than ASCs; however, the apoptosis and immunomodulatory marker expression was similar in both populations. Native ASCs and HPCs exhibited different baseline expressions of markers involved in mitochondrial dynamics, the oxidative stress response, apoptosis and inflammation. When exposed to a pro-inflammatory microenvironment, ASCs and HPCs differed in the expression of mitochondrial condition markers and chosen miRNAs.

Although mesenchymal stem cells (MSCs) can be obtained from various tissues such as bone marrow (BM), umbilical cord blood (UCB) and umbilical cord tissue (UC), the comparative efficacy of each MSC in tendon regeneration is unknown. Therefore, we investigated the efficacy of MSCs isolated from three different sources on tendon regeneration after injury. We evaluated the potential of BM-, UCB- and UC-MSC to differentiate into tendon-like cells in tensioned three-dimensional construct (T-3D) using gene and histological analysis. In animal experiments, full-thickness tendon defect (FTD) was created in supraspinatus of rats, and injected with Saline and BM-, UCB- and UC-MSC. After two and four weeks, histological evaluations were performed. After inducing tenogenic differentiation, the gene expression of scleraxis, mohawk, type I collagen and tenascin-C was upregulated by 3.12-, 5.92-, 6.01- and 1.61-fold respectively and formation of tendon-like matrix was increased 4.22-fold in UC-MSC compared to BM-MSC in T-3D. In animal experiments, the total degeneration score was lower in the UC-MSC group than in BM-MSC group at both weeks. In heterotopic matrix formation, glycosaminoglycan-rich area was reduced in the UC-MSC group, whereas area was larger in the BM-MSC group than in Saline group at four weeks. In conclusion, UC-MSC is superior to other MSCs in differentiating into tendon-like lineage cells and forming a well-organized tendon-like matrix under T-3D conditions. UC-MSC enhances regeneration of FTD in terms of histological properties compared to BM- and UCB-MSC.

Accumulating evidence indicates that most primary Wharton's jelly mesenchymal stem cells (WJ-MSCs) therapeutic potential is due to their paracrine activity, i.e., their ability to modulate their microenvironment by releasing bioactive molecules and factors collectively known as secretome. These bioactive molecules and factors can either be released directly into the surrounding microenvironment or can be embedded within the membrane-bound extracellular bioactive nano-sized (usually 30-150 nm) messenger particles or vesicles of endosomal origin with specific route of biogenesis, known as exosomes or carried by relatively larger particles (100 nm-1 μm) formed by outward blebbing of plasma membrane called microvesicles (MVs); exosomes and MVs are collectively known as extracellular vesicles (EVs). The bioactive molecules and factors found in secretome are of various types, including cytokines, chemokines, cytoskeletal proteins, integrins, growth factors, angiogenic mediators, hormones, metabolites, and regulatory nucleic acid molecules. As expected, the secretome performs different biological functions, such as immunomodulation, tissue replenishment, cellular homeostasis, besides possessing anti-inflammatory and anti-fibrotic effects. This review highlights the current advances in research on the WJ-MSCs' secretome and its prospective clinical applications.

During the pandemic of severe respiratory distress syndrome coronavirus 2 (SARS-CoV2), many novel therapeutic modalities to treat Coronavirus 2019 induced disease (COVID-19) were explored. This study summarizes 195 clinical trials of advanced cell therapies targeting COVID-19 that were registered over the two years between January 2020 to December 2021. In addition, this work also analyzed the cell manufacturing and clinical delivery experience of 26 trials that published their outcomes by July 2022. Our demographic analysis found the highest number of cell therapy trials for COVID-19 was in United States, China, and Iran (N=53, 43, and 19, respectively), with the highest number per capita in Israel, Spain, Iran, Australia, and Sweden (N=0.641, 0.232, 0,223, 0.194, and 0.192 trials per million inhabitants). The leading cell types were multipotent mesenchymal stromal/stem cells (MSCs), natural killer (NK) cells, and mononuclear cells (MNCs), accounting for 72%, 9%, and 6% of the studies, respectively. There were 24 published clinical trials that reported on infusions of MSCs. A pooled analysis of these MSC studies found that MSCs provide a relative risk reduction for all-cause COVID-19 mortality of RR=0.63 (95% CI 0.46 to 0.85). This result corroborates previously published smaller meta-analyses, which suggested that MSC therapy demonstrated a clinical benefit for COVID-19 patients. The sources of the MSCs used in these studies and their manufacturing and clinical delivery methods were remarkably heterogeneous, with some predominance of perinatal tissue-derived products. Our results highlight the important role that cell therapy products may play as an adjunct therapy in the management of COVID-19 and its related complications, as well as the importance of controlling key manufacturing parameters to ensure comparability between studies. Thus, we support ongoing calls for a global registry of clinical studies with MSC products that could better link cell product manufacturing and delivery methods to clinical outcomes. Although advanced cell therapies may provide an important adjunct treatment for patients affected by COVID-19 in the near future, preventing pathology through vaccination still remains the best protection to date. We conducted a systematic review and meta-analysis of advanced cell therapy clinical trials as potential novel treatment for COVID-19 (resulting from SARS-CoV-2 coronavirus infection), including analysis of the global clinical trial landscape, published safety/efficacy outcomes (RR/OR), and details on cell product manufacturing and clinical delivery. This study had a 2-year observation interval from start of January 2020 to end of December 2021, including a follow-up period until end of July to identify published outcomes, which covers the most vivid period of clinical trial activity, and is also the longest observation period studied until today. In total, we identified 195 registered advanced cell therapy studies for COVID-19, employing 204 individual cell products. Leading registered trial activity was attributed to the USA, China, and Iran. Through the end of July 2022, 26 clinical trials were published, with 24 out of 26 articles employing intravenous infusions (IV) of mesenchymal stromal/stem cell (MSC) products. Most of the published trials were attributed to China and Iran. The cumulative results from the 24 published studies employing infusions of MSCs indicated an improved survival (RR=0.63 with 95% Confidence Interval 0.46 to 0.85). Our study is the most comprehensive systematic review and meta-analysis on cell therapy trials for COVID-19 conducted to date, clearly identifying the USA, China, and Iran as leading advanced cell therapy trial countries for COVID-19, with further strong contributions from Israel, Spain, Australia and Sweden. Although advanced cell therapies may provide an important adjunct treatment for patients affected by COVID-19 in the future, preventing pathology through vaccination remains the best protection.

Atopic dermatitis (AD) is one of the most common immune and inflammatory skin disorders, leading to insufferable itching and skin abnormalities that seriously affect life quality of patients. There are still huge unmet needs for long-term and effective disease control, despite currently available therapies. Evidenced by some preclinical and clinical studies of AD treatment with stem cells, stem cell treatment could significantly and effectively ameliorate AD symptoms.

To elucidate underlying mechanisms of how stem cells therapy alleviates AD-like symptoms.

An AD-like mouse model was constructed and treated with mesenchymal stem cells (MSCs) subcutaneously or subcutaneously combined with intravenously. The differentially expressed genes were sorted out from RNA sequencing results of dorsal skin and blood.

Two injection routes of MSCs could alleviate AD-like symptoms and pathologic changes of the skin and immune organs. RNA sequencing of dorsal skin sections and blood provided gene expression signatures for amelioration of skin defects, inflammatory and immune modulation by MSCs, as well as common AD molecular markers for the skin and blood, which may benefit for clinical diagnosis. IL-1β and its signaling pathway were specifically found to be associated with the development of AD-like dermatitis lesions. MSC treatment effectively inhibited the JAK-STAT pathway and receptors of IL-4, IL-13, IL-17, and IgE.

MSC therapy could regulate abnormal immune and inflammatory status in AD. Mechanistic exploration will contribute to the development of personalized AD treatment based on MSCs.

Umbilical cord mesenchymal stem cells (UCMSCs) are a reportedly promising choice in the treatment of irreversible pulmonary fibrosis and lethal interstitial lung disease with limited drug treatment options. In this study, we investigated the therapeutic efficacy of UCMSCs overexpressing hepatocyte growth factor (HGF), which is considered one of the main anti-fibrotic factors secreted by MSCs. Adenovirus vector carrying the HGF gene was transfected into UCMSCs to produce HGF-modified UCMSCs (HGF-UCMSCs). Transfection promoted the proliferation of UCMSCs and did not change the morphology, and differentiation ability, or biomarkers. Rats were injected with HGF-UCMSCs on days 7 and 11 after intratracheal administration of bleomycin (10 mg/kg). We performed an analysis of histopathology and lung function to evaluate the anti-fibrotic effect. The results showed that HGF-UCMSCs decreased the Ashcroft scores in hematoxylin and eosin-stained sections, the percentage positive area in Masson trichrome-stained sections, and the hydroxyproline level in lungs. Forced expiratory volume in the first 300 m/forced vital capacity was also improved by HGF-UCMSCs. To explore the possible therapeutic mechanism of HGF-UCMSCs, we detected inflammatory factors in the lungs and performed mRNA sequencing in UCMSCs and HGF-UCMSCs. The data indicated that inhibition of interleukin-17 in the lung may be related to the anti-fibrosis of HGF-UCMSCs, and overexpressed HGF probably played a primary role in the treatment. Collectively, our study findings suggested that the overexpression of HGF may improve the anti-fibrotic effect of UCMSCs through directly or indirectly interacting with interleukin-17-producing cells in fibrotic lungs.

Mesenchymal stem cells (MSCs) are pluripotent stem cells with multidirectional differentiation potential and strong immunomodulatory capacity. MSCs have been widely used in the treatment of injured, inflammatory, and immune-related diseases. Resting MSCs lack differentiation and immunomodulatory ability. Instead, they rely on microenvironmental factors to: 1) stimulate and regulate their expression of specific cell growth factors, chemokines, immunomodulatory factors, or receptors; or 2) direct their differentiation into specific tissue cells, which ultimately perform tissue regeneration and repair and immunomodulatory functions. Tumor necrosis factor (TNF)-α is central to the creation of an inflammatory microenvironment. TNF-α regulates the fate and functional reprogramming of MSCs, either alone or in combination with a variety of other inflammatory factors. TNF-α can exert opposing effects on MSCs, from inducing MSC apoptosis to enhancing their anti-tumor capacity. In addition, the immunomodulation and osteogenic differentiation capacities of MSCs, as well as their exosome or microvesicle components vary significantly with TNF-α stimulating concentration, time of administration, or its use in combination with or without other factors. Therefore, this review discusses the impact of TNF-α on the fate and functional reprogramming of MSCs in the inflammatory microenvironment, to provide new directions for improving the immunomodulatory and tissue repair functions of MSCs and enhance their therapeutic potential.

It is often difficult to regain neurological function following spinal cord injury (SCI). Neuroinflammation is thought to be responsible for this failure. Regulating the inflammatory response post-SCI may contribute to the recovery of neurological function. Over the past few decades, studies have found that macrophages/microglia are one of the primary effector cells in the inflammatory response following SCI. Growing evidence has documented that macrophages/microglia are plastic cells that can polarize in response to microenvironmental signals into M1 and M2 macrophages/microglia. M1 produces pro-inflammatory cytokines to induce inflammation and worsen tissue damage, while M2 has anti-inflammatory activities in wound healing and tissue regeneration. Recent studies have indicated that the transition from the M1 to the M2 phenotype of macrophage/microglia supports the regression of inflammation and tissue repair. Here, we will review the role of the inflammatory response and macrophages/microglia in SCI and repair. In addition, we will discuss potential molecular mechanisms that induce macrophage/microglia polarization, with emphasis on neuroprotective therapies that modulate macrophage/microglia polarization, which will provide new insights into therapeutic strategies for SCI.

Sepsis is a debilitating systemic inflammation that resulted from infection or injury. Despite many advances in treatment, the resulting mortality rate has remained high due to increasing antibiotic resistance and aging communities. The present study investigated the effects of stem cell-derived exosomes in a mouse model of LPS-induced systemic inflammation.

To induce sepsis, the LPS model was used. Mice were divided into three groups: normal, patient group (LPS + PBS), and treatment group (LPS + exosome). The treatment group received an intravenous exosome 1 h after induction of the model. Patient and treatment groups were sacrificed at 4, 6, 24, and 48 h after induction of the model, and their tissues were isolated. Blood samples were taken from animal hearts to perform biochemical and immunological tests. The study results were analyzed using Graph Pad Prism software version 9.

Mesenchymal stem cell-derived exosomes decreased serum levels of ALT and AST liver enzymes, decreased neutrophil to lymphocyte ratio (NLR), and improved kidney, liver, and lung tissue damage at 4, 6, and 24 h after model induction. At 24 h, the exosomes were able to reduce serum urea levels. This study revealed decreased levels of inflammatory cytokines such as IL-6, IL-1β, and TNF-α after exosome injection.

Our findings suggest that treating mice with stem cell-derived exosomes can ameliorate the destructive effects of inflammation caused by sepsis by reducing inflammatory factors and tissue damage.

Three-dimensional (3D) culture of human mesenchymal stem cells (MSCs) as spheroids enhances the production of important regulators of inflammation: prostaglandin E2 (PGE₂), interleukin (IL)-6, and tumor necrosis factor-inducible gene 6 (TSG-6). The horse is a model species and suffers from musculoskeletal, ocular, and systemic inflammatory disease. It is unknown if 3D culture promotes enhanced production of immunomodulatory cytokines and regulators in equine MSCs and if there is variation between individual cell donors. We evaluated the feasibility, cell viability, and stem cell marker stability of 3D-cultured equine bone marrow-derived MSCs (eBMSCs) and determined the effect of inflammatory stimulation upon gene expression and secretion of key regulators of inflammation [PGE₂, TSG-6, IL-10, IL-6, stromal cell-derived factor 1 (SDF-1)]. Variations in anti-inflammatory phenotype between six donors were investigated, with and without IL-1β stimulation, in either monolayer [two-dimensional (2D)] or 3D culture. Our results showed that eBMSCs self-aggregate in 3D culture while maintaining cell viability and markers of stemness CD90, CD44, CD104, and Oct4. In addition, 3D culture enhances the anti-inflammatory phenotype regardless of inflammatory stimulation by increasing PGE₂, IL-6, TSG-6, SDF-1, and IL-10. Finally, anti-inflammatory phenotype was enhanced by IL-1β exposure but showed significant variation between cell lines in the degree of gene upregulation, and what genes were expressed. We conclude that 3D culture of eBMSCs as spheroids alters their anti-inflammatory phenotype, but this effect is influenced by cytokine exposure and cell donor.

Mesenchymal stromal cells (MSCs) show immunosuppressive effects both via cell-to-cell contact (direct) with immune cells and by producing paracrine factors and extracellular vesicles (indirect). A key challenge in delivering this therapeutic effect in vivo is retaining the MSCs at the site of injection. One way to address this is by encapsulating the MSCs within suitable biomaterial scaffolds. Here, we assess the immunosuppressive effect of alginate-encapsulated murine MSCs on proliferating murine splenocytes. Our results show that MSCs are able to significantly suppress splenocyte proliferation by ∼50% via the indirect mechanism and almost completely (∼98%) via the direct mechanism. We also show for the first time that MSCs as monolayers on tissue culture plastic or encapsulated within alginate, when physically isolated from the splenocytes via transwells, are able to sustain immunosuppressive activity with repeated exposure to fresh splenocytes, for as long as 9 days. These results indicate the need to identify design strategies to simultaneously deliver both modes of MSC immunosuppression. By designing cell-biomaterial constructs with tailored degradation profiles, we can achieve a more sustained (avoiding MSCs migration and apoptosis) and controlled release of both the paracrine signals and eventually the cells themselves enabling efficient MSC-based immunosuppressive therapies for wound healing.

Inflammatory processes contribute to osteoarthritis (OA) severity and progression. Mesenchymal stem cells, particularly those derived from adipose tissue (ASCs), are able to sense and control the inflammatory environment. This immunomodulatory potential can be boosted by different priming strategies based on inflammatory stimulation. The aim of the present study is to investigate the transcriptional modulation of a huge panel of genes and functionally verify the predicted immunomodulatory ability of ASCs after interleukin one beta (IL-1β) priming. ASCs were isolated from adipose tissue obtained from three donors and expanded. After stimulation with 1 ng/ml of IL-1β for 48 h, cells were collected for gene array and functional tests. Pooled cells from three donors were used for RNA extraction and gene array analysis. Gene Ontology (GO) enrichment analysis and Gene Set Enrichment Analysis (GSEA) were performed to assess the involvement of the modulated genes after priming in specific biological processes and pathways. Functional co-culture tests of ASCs with T cells and macrophages were performed to assess the ability of primed ASCs to modulate immune cell phenotype. Among the overall genes analyzed in the gene array, about the 18% were up- or down-regulated in ASCs after IL-1β priming. GO enrichment analysis of up- or down-regulated genes in ASCs after IL-1β priming allowed identifying specific pathways involved in the modulation of inflammation and extracellular matrix remodeling. The main processes enriched according to the GSEA are related to the inflammatory response and cell proliferative processes. Functional tests on immune cells showed that primed and non-primed ASCs induced a decrease in the CD3+ T lymphocytes survival rate and an anti-inflammatory macrophage polarization. In conclusion, IL-1β priming represents a tailored strategy to enhance the ability of ASCs to direct macrophages towards an anti-inflammatory phenotype and, consequently, improve the efficacy of ASCs in counteracting the OA inflammatory component.

Immune cells play an important role in the development of inflammation in type 1 diabetes mellitus, so we want to explore the changes of CD4+ T cells and macrophages in vivo, which can provide an experimental basis for immunotherapy based on CD4+ T cells and macrophages. The intraperitoneal injection of streptozocin was used to induce a type 1 diabetes mellitus mouse model; the blood glucose, body weight, and the expression of inflammatory factors in the kidney were measured. Immunohistochemistry was applied to determine and analyze the infiltration of CD4+ T cells and macrophages in the spleen, pancreas, and kidney. The subtypes of macrophages in the kidney and CD4+ T cells in the spleen were analyzed by flow cytometry. Our study suggests that CD4+ T cells and macrophages increase, while the inflammatory immune response system is activated in the development of T1DM. CD4+ T cells positively correlated with macrophages in the pancreas and kidney of T1DM. CD4+ T cells turn to pro-inflammatory subtypes in the spleen of T1DM, while macrophages turn to pro-inflammatory subtypes in the kidney of T1DM. Therefore, regulation of CD4+ T cells and macrophages may be a potential target for T1DM and kidney complications.

Numerous studies have shown that mesenchymal stromal cells (MSCs) promote cutaneous wound healing via paracrine signaling. Our previous study found that the secretome of MSCs was significantly amplified by treatment with IFN-γ and TNF-α (IT). It has been known that macrophages are involved in the initiation and termination of inflammation, secretion of growth factors, phagocytosis, cell proliferation, and collagen deposition in wound, which is the key factor during wound healing. In this study, we aim to test whether the supernatant of MSCs pretreated with IT (S-IT MSCs) possesses a more pronounced effect on improving wound healing and describe the interplay between S-IT MSCs and macrophages as well as the potential mechanism in skin wound healing.

In the present study, we used a unique supernatant of MSCs from human umbilical cord-derived MSCs (UC-MSCs) pretreated with IT, designated S-IT MSCs, subcutaneously injected into a mice total skin excision. We evaluated the effect of S-IT MSCs on the speed and quality of wound repair via IT MSCs-derived IL-6-dependent M2 polarization in vivo by hematoxylin-eosin staining (H&E), immunohistochemistry (IHC), immunofluorescence (IF), Masson's trichrome staining, Sirius red staining, quantitative real-time PCR (qPCR). In addition, the effect of S-IT MSCs on the polarization of macrophages toward M2 phenotype and the potential mechanism of it were also investigated in vitro by flow cytometry (FCM), enzyme-linked immunosorbent assay (ELISA), tube formation assay, and western blot analysis.

Compared with control supernatant (S-MSCs), our H&E and IF results showed that S-IT MSCs were more effectively in promoting macrophages convert to the M2 phenotype and enhancing phagocytosis of M2 macrophages. Meanwhile, the results of tube formation assay, IHC, Masson's trichrome staining, Sirius red staining showed that the abilities of M2 phenotype to promote vascularization and collagen deposition were significantly enhanced by S-IT MSCs-treated, thereby accelerating higher quality wound healing. Further, our ELISA, FCM, qPCR and western blot results showed that IL-6 was highly enriched in S-IT MSCs and acted as a key regulator to induce macrophages convert to the M2 phenotype through IL-6-dependent signaling pathways, ultimately achieving the above function of promoting wound repair.

These findings provide the first evidence that the S-IT MSCs is more capable of eliciting M2 polarization of macrophages via IL-6-dependent signaling pathways and accelerating wound healing, which may represent a new strategy for optimizing the therapeutic effect of MSCs on wound healing.