Tensile force impairs lip muscle regeneration under the regulation of interleukin-10.

BACKGROUND: Orbicularis oris muscle, the crucial muscle in speaking, facial expression and aesthetics, is considered the driving force for optimal lip repair. Impaired muscle regeneration remains the main culprit for unsatisfactory surgical outcomes. However, there is a lack of study on how different surgical manipulations affect lip muscle regeneration, limiting efforts to seek effective interventions. METHODS: In this study, we established a rat lip surgery model where the orbicularis oris muscle was injured by manipulations including dissection, transection and stretch. The effect of each technique on muscle regeneration was examined by histological analysis of myogenesis and fibrogenesis. The impact of tensile force was further investigated by the in vitro application of mechanical strain on cultured myoblasts. Transcriptome profiling of muscle satellite cells from different surgical groups was performed to figure out the key factors mediating muscle fibrosis, followed by therapeutic intervention to improve muscle regeneration after lip surgeries. RESULTS: Evaluation of lip muscle regeneration till 56 days after injury revealed that the stretch group resulted in the most severe muscle fibrosis (n = 6, fibrotic area 48.9% in the stretch group, P < 0.001, and 25.1% in the dissection group, P < 0.001). There was the lowest number of Pax7-positive nuclei at Days 3 and 7 in the stretch group (n = 6, P < 0.001, P < 0.001), indicating impaired satellite cell expansion. Myogenesis was impaired in both the transection and stretch groups, as evidenced by the delayed peak of centrally nucleated myofibers and embryonic MyHC. Meanwhile, the stretch group had the highest percentage of Pdgfra+ fibro-adipogenic progenitors infiltrated area at Days 3, 7 and 14 (n = 6, P = 0.003, P = 0.006, P = 0.037). Cultured rat lip muscle myoblasts exhibited impaired myotube formation and fusion capacity when exposed to a high magnitude (ε = 2688 µ strain) of mechanical strain (n = 3, P = 0.014, P = 0.023). RNA-seq analysis of satellite cells isolated from different surgical groups demonstrated that interleukin-10 was the key regulator in muscle fibrosis. Administration of recombinant human Wnt7a, which can inhibit the expression of interleukin-10 in cultured satellite cells (n = 3, P = 0.041), exerted an ameliorating effect on orbicularis oris muscle fibrosis after stretching injury in surgical lip repair. CONCLUSIONS: Tensile force proved to be the most detrimental manoeuvre for post-operative lip muscle regeneration, despite its critical role in correcting lip and nose deformities. Adjunctive biotherapies to regulate the interleukin-10-mediated inflammatory process could facilitate lip muscle regeneration under conditions of high surgical tensile force.

Human pancreatic islet-derived stromal cells reveal combined features of mesenchymal stromal cells and pancreatic stellate cells.

BACKGROUND: Mesenchymal stromal cells (MSCs) are recognized for their potential in regenerative medicine, attributed to their multipotent differentiation capabilities and immunomodulatory properties. Despite this potential, the classification and detailed characterization of MSCs, especially those derived from specific tissues like the pancreas, remains challenging leading to a proliferation of terminology in the literature. This study aims to address these challenges by providing a thorough characterization of human pancreatic islets-derived mesenchymal stromal cells (hPD-MSCs). METHODS: hPD-MSCs were isolated from donor islets using enzymatic digestion, immortalized through lentiviral transduction of human telomerase reverse transcriptase (hTERT). Cells were characterized by immunostaining, flow cytometry and multilineage differentiation potential into adipogenic and osteogenic lineages. Further a transcriptomic analysis was done to compare the gene expression profiles of hPD-MSCs with other mesenchymal cells. RESULTS: We show that hPD-MSCs express the classical MSC features, including morphological characteristics, surface markers expression (CD90, CD73, CD105, CD44, and CD106) and the ability to differentiate into both adipogenic and osteogenic lineages. Furthermore, transcriptomic analysis revealed distinct gene expression profiles, showing notable similarities between hPD-MSCs and pancreatic stellate cells (PSCs). The study also identified specific genes that distinguish hPD-MSCs from MSCs of other origins, including genes associated with pancreatic function (e.g., ISL1) and neural development (e.g., NPTX1, ZNF804A). A novel gene with an unknown function (ENSG00000286190) was also discovered. CONCLUSIONS: This study enhances the understanding of hPD-MSCs, demonstrating their unique characteristics and potential applications in therapeutic strategies. The identification of specific gene expression profiles differentiates hPD-MSCs from other mesenchymal cells and opens new avenues for research into their role in pancreatic function and neural development.

1,25-dihydroxyvitamin-D3 distinctly impacts the paracrine and cell-to-cell contact interactions between hPDL-MSCs and CD4+ T lymphocytes.

Introduction: Human periodontal ligament-derived mesenchymal stromal cells (hPDL-MSCs) possess a strong ability to modulate the immune response, executed via cytokine-boosted paracrine and direct cell-to-cell contact mechanisms. This reciprocal interaction between immune cells and hPDL-MSCs is influenced by 1,25-dihydroxyvitamin-D3 (1,25(OH)2D3). In this study, the participation of different immunomodulatory mechanisms on the hPDL-MSCs-based effects of 1,25(OH)2D3 on CD4+ T lymphocytes will be elucidated using different co-culture models with various cytokine milieus. Material and methods: hPDL-MSCs and CD4+ T lymphocytes were co-cultured indirectly and directly with inserts (paracrine interaction only) or directly without inserts (paracrine and direct cell-to-cell contact interaction). They were stimulated with TNF-α or IL-1ß in the absence/presence of 1,25(OH)2D3. After five days of co-cultivation, the CD4+ T lymphocyte proliferation, viability, and cytokine secretion were analyzed. Additionally, the gene expression of soluble and membrane-bound immunomediators was determined in hPDL-MSCs. Results: In the indirect and direct co-culture model with inserts, 1,25(OH)2D3 decreased CD4+ T lymphocyte proliferation and viability. The direct co-culture model without inserts caused the opposite effect. 1,25(OH)2D3 mainly decreased the CD4+ T lymphocyte-associated secretion of cytokines via hPDL-MSCs. The degree of these inhibitions varied between the different co-culture setups. 1,25(OH)2D3 predominantly decreased the expression of the soluble and membrane-bound immunomediators in hPDL-MSCs to a different extent, depending on the co-culture models. The degree of all these effects depended on the absence and presence of exogenous TNF-α and IL-1ß. Conclusion: These data assume that 1,25(OH)2D3 differently affects CD4+ T lymphocytes via the paracrine and direct cell-to-cell contact mechanisms of hPDL-MSCs, showing anti- or pro-inflammatory effects depending on the co-culture model type. The local cytokine microenvironment seems to be involved in fine-tuning these effects. Future studies should consider this double-edged observation by executing different co-culture models in parallel.

Distinct muscle regenerative capacity of human induced pluripotent stem cell-derived mesenchymal stromal cells in Ullrich congenital muscular dystrophy model mice.

BACKGROUND: Ullrich congenital muscular dystrophy (UCMD) is caused by a deficiency in type 6 collagen (COL6) due to mutations in COL6A1, COL6A2, or COL6A3. COL6 deficiency alters the extracellular matrix structure and biomechanical properties, leading to mitochondrial defects and impaired muscle regeneration. Therefore, mesenchymal stromal cells (MSCs) that secrete COL6 have attracted attention as potential therapeutic targets. Various tissue-derived MSCs exert therapeutic effects in various diseases. However, no reports have compared the effects of MSCs of different origins on UCMD pathology. METHODS: To evaluate which MSC population has the highest therapeutic efficacy for UCMD, in vivo (transplantation of MSCs to Col6a1-KO/NSG mice) and in vitro experiments (muscle stem cell [MuSCs] co-culture with MSCs) were conducted using adipose tissue-derived MSCs, bone marrow-derived MSCs, and xeno-free-induced iPSC-derived MSCs (XF-iMSCs). RESULTS: In transplantation experiments on Col6a1-KO/NSG mice, the group transplanted with XF-iMSCs showed significantly enhanced muscle fiber regeneration compared to the other groups 1 week after transplantation. At 12 weeks after transplantation, only the XF-iMSCs transplantation group showed a significantly larger muscle fiber diameter than the other groups without inducing fibrosis, which was observed in the other transplantation groups. Similarly, in co-culture experiments, XF-iMSCs were found to more effectively promote the fusion and differentiation of MuSCs derived from Col6a1-KO/NSG mice than the other primary MSCs investigated in this study. Additionally, in vitro knockdown and supplementation experiments suggested that the IGF2 secreted by XF-iMSCs promoted MuSC differentiation. CONCLUSION: XF-iMSCs are promising candidates for promoting muscle regeneration while avoiding fibrosis, offering a safer and more effective therapeutic approach for UCMD than other potential therapies.

The influence of fetal bovine serum concentration on stemness and neuronal differentiation markers in stem cells from human exfoliated deciduous teeth.

Dental Stem Cells (DSCs) from discarded teeth are a non-invasive and ethically favorable source with the potential for neurogenesis due to their ectodermal origin. Stem cells from human exfoliated deciduous teeth (SHED) are particularly promising due to their high differentiation potential and relative immaturity compared to other Mesenchymal Stromal Cells (MSCs). Markers like CD56 and CD271 are critical in identifying MSC subpopulations for therapeutic applications because of their roles in neurodevelopment and maintaining stemness. This study investigates how fetal bovine serum (FBS) concentrations affect the expression of CD56 and CD271 in SHED, influencing their stemness and neuronal differentiation potential. SHEDs were isolated from various donors, cultured, and characterized for MSC traits using markers such as CD14, CD19, CD29, CD34, CD45, CD73, CD90, CD105, CD56, and CD271. Culturing SHED in different FBS conditions (standard 15 %, reduced 1 % and 5 %, and FBS-free) showed that lower FBS concentrations increase CD271 and CD56 expression while maintaining the standard MSC immunophenotype. Importantly, the enhanced expression of these markers can be induced even after SHEDs have been expanded in standard FBS concentrations. These findings suggest that FBS concentration can optimize SHED culture conditions, enhancing their suitability for regenerative medicine and tissue engineering applications.

The Therapeutic Potential of Exosome Therapy in Sepsis Management: Addressing Complications and Improving Outcomes".

Infection occurs when pathogens penetrate tissues, reproduce, and trigger a host response to both the infectious agents and their toxins. A diverse array of pathogens, including viruses and bacteria, can cause infections. The host's immune system employs several mechanisms to combat these infections, typically involving an innate inflammatory response. Inflammation is a complex biological reaction that can affect various parts of the body and is a key component of the response to harmful stimuli. Sepsis arises when the body's response to infection leads to widespread damage to tissues and organs, potentially resulting in severe outcomes or death. The initial phase of sepsis involves immune system suppression. Early identification and targeted management are crucial for improving sepsis outcomes. Common treatment approaches include antibiotics, intravenous fluids, blood cultures, and monitoring urine output. This study explores the potential of exosome therapy in enhancing the management and alleviation of sepsis symptoms.

Chondrogenic Potential of Umbilical Cord-Derived Mesenchymal Stromal Cells: Insights and Innovations.

Background: The advent of tissue engineering and regenerative medicine has introduced innovative approaches to treating degenerative and traumatic injuries, particularly in cartilage, a tissue with limited self-repair capabilities. Among the various stem cell sources, umbilical cord-derived mesenchymal stromal cells (UC-MSCs) have garnered significant interest due to their non-invasive collection, minimal ethical concerns, and robust regenerative potential, particularly in cartilage regeneration. Methods: A comprehensive literature review was conducted using multiple databases, including PubMed, Scopus, Web of Science, and Google Scholar. Search terms focused on "umbilical cordderived mesenchymal stromal cells," "chondrogenesis," "cartilage regeneration," and related topics. Studies published in the past two decades were included, with selection criteria emphasizing methodological rigor and relevance to UC-MSC chondrogenesis. The review synthesizes findings from various sources to provide a thorough analysis of the potential of UC-MSCs in cartilage tissue engineering. Results: UC-MSCs exhibit significant chondrogenic potential, supported by their ability to differentiate into chondrocytes under specific conditions. Recent advancements include the development of biomaterial scaffolds and the application of genetic engineering techniques, such as CRISPR/Cas9, to enhance chondrogenic differentiation. Despite these advancements, challenges remain in standardizing cell isolation techniques, scaling up production for clinical use, and ensuring the long-term functionality of regenerated cartilage. Conclusion: UC-MSCs offer a promising solution for cartilage regeneration in the field of regenerative medicine. Ongoing research is focused on overcoming current challenges through the use of advanced technologies, including bioreactors and gene editing. Collaborative efforts among researchers, clinicians, and bioengineers are essential to translating the potential of UC-MSCs into effective clinical therapies, which could significantly advance tissue regeneration and therapeutic innovation.

Understanding and controlling the variables for stromal vascular fraction therapy.

In regenerative medicine, the isolation of mesenchymal stromal cells (MSCs) from the adipose tissue's stromal vascular fraction (SVF) is a critical area of study. Our review meticulously examines the isolation process of MSCs, starting with the extraction of adipose tissue. The choice of liposuction technique, anatomical site, and immediate processing are essential to maintain cell functionality. We delve into the intricacies of enzymatic digestion, emphasizing the fine-tuning of enzyme concentrations to maximize cell yield while preventing harm. The review then outlines the filtration and centrifugation techniques necessary for isolating a purified SVF, alongside cell viability assessments like flow cytometry, which are vital for confirming the efficacy of the isolated MSCs. We discuss the advantages and drawbacks of using autologous vs allogeneic SVF sources, touching upon immunocompatibility and logistical considerations, as well as the variability inherent in donor-derived cells. Anesthesia choices, the selection between hypodermic needles vs liposuction cannulas, and the role of adipose tissue lysers in achieving cellular dissociation are evaluated for their impact on SVF isolation. Centrifugation protocols are also analyzed for their part in ensuring the integrity of the SVF. The necessity for standardized MSC isolation protocols is highlighted, promoting reproducibility and successful clinical application. We encourage ongoing research to deepen the understanding of MSC biology and therapeutic action, aiming to further the field of regenerative medicine. The review concludes with a call for rigorous research, interdisciplinary collaboration, and strict adherence to ethical and regulatory standards to safeguard patient safety and optimize treatment outcomes with MSCs.

Microvesicles derived from mesenchymal stem cells inhibit acute respiratory distress syndrome-related pulmonary fibrosis in mouse partly through hepatocyte growth factor.

BACKGROUND: Pulmonary fibrosis is one of the main reasons for the high mortality rate among acute respiratory distress syndrome (ARDS) patients. Mesenchymal stromal cell-derived microvesicles (MSC-MVs) have been shown to exert antifibrotic effects in lung diseases. AIM: To investigate the effects and mechanisms of MSC-MVs on pulmonary fibrosis in ARDS mouse models. METHODS: MSC-MVs with low hepatocyte growth factor (HGF) expression (siHGF-MSC-MVs) were obtained via lentivirus transfection and used to establish the ARDS pulmonary fibrosis mouse model. Following intubation, respiratory mechanics-related indicators were measured via an experimental small animal lung function tester. Homing of MSC-MVs in lung tissues was investigated by near-infrared live imaging. Immunohistochemical, western blotting, ELISA and other methods were used to detect expression of pulmonary fibrosis-related proteins and to compare effects on pulmonary fibrosis and fibrosis-related indicators. RESULTS: The MSC-MVs gradually migrated and homed to damaged lung tissues in the ARDS model mice. Treatment with MSC-MVs significantly reduced lung injury and pulmonary fibrosis scores. However, low expression of HGF (siHGF-MSC-MVs) significantly inhibited the effects of MSC-MVs (P < 0.05). Compared with the ARDS pulmonary fibrosis group, the MSC-MVs group exhibited suppressed expression of type I collagen antigen, type III collagen antigen, and the proteins transforming growth factor-ß and α-smooth muscle actin, whereas the siHGF-MVs group exhibited significantly increased expression of these proteins. In addition, pulmonary compliance and the pressure of oxygen/oxygen inhalation ratio were significantly lower in the MSC-MVs group, and the effects of the MSC-MVs were significantly inhibited by low HGF expression (all P < 0.05). CONCLUSION: MSC-MVs improved lung ventilation functions and inhibited pulmonary fibrosis in ARDS mice partly via HGF mRNA transfer.

Tissue-specific functions of MSCs are linked to homeostatic muscle maintenance and alter with aging.

Mesenchymal stromal cells (MSCs), also known as fibro-adipogenic progenitors, play a critical role in muscle maintenance and sarcopenia development. Although analogous MSCs are present in various tissues, recent single-cell RNA-seq studies have revealed the inter-tissue heterogeneity of MSCs. However, the functional significance of MSC heterogeneity and its role in aging remain unclear. Here, we investigated the properties of MSCs and their age-related changes in seven mouse tissues through histological, cell culture, and genetic examinations. The tissue of origin had a greater impact on the MSC transcriptome than aging. By first analyzing age-related changes, we found that Kera is exclusively expressed in muscle MSCs and significantly down-regulated by aging. Kera knockout mice recapitulated some sarcopenic phenotypes including reduced muscle mass and specific force, revealing the functional importance of Kera in the maintenance of muscle youth. These results suggest that MSCs have tissue-specific supportive functions and that deterioration in these functions may trigger tissue aging.

Innovative cell therapies for systemic sclerosis: available evidence and new perspectives.

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

Quality Assessment of Long-Term Cryopreserved Human Bone-Derived Marrow Mesenchymal Stromal Cell Samples: Experience from the Texas Heart Institute Biorepository and Biospecimen Profiling Core.

In biomedical research, biorepositories are pivotal resources that safeguard and supply clinical samples for scientific investigators. Proper long-term cryopreservation conditions are essential to maintain biospecimen quality. In this study, we analyzed the efficacy of sample cryopreservation at the Texas Heart Institute Biorepository and Biospecimen Profiling Core (THI-BRC). Our assessments included a thorough review of internal processes, quality reports, and both internal and external audit outcomes. We examined the integrity of human bone marrow-derived multipotent mesenchymal stromal cells (BM-MSCs) that were cryopreserved for over 5 years. These samples originated from randomly selected clinical trial participants or commercially sourced cell lines. Parameters such as cell viability, DNA and RNA integrity, population doubling time, sterility, and BM-MSC-specific attributes such as surface antigen expression and differentiation potential were studied. BM-MSC samples cryopreserved for ∼6 months served as our control. Our results demonstrated that the 5-year cryopreserved samples maintained their integrity compared with the shorter-term stored control samples. Moreover, THI-BRC has met accreditation agency standards and has not received any repeated deficiencies over 7 years. Collectively, our findings affirm that THI-BRC's biospecimen storage protocols align with accepted standards as confirmed by the quality assessment of long-term stored clinical samples.

[Prerequisite and organisation of health-care pathways for Cell and Gene therapies, using Mesenchymal Stromal Cells (MSC) or Chimeric Antigen Receptor (CAR) T cells, in patients with autoimmune systemic diseases]. / Prérequis et organisation du parcours de soins en vue de l'utilisation d'une thérapie cellulaire ou génique par cellules stromales mésenchymateuses (CSM) ou par cellules T porteuses d'un récepteur antigénique chimérique (CAR-T cells) chez les patients avec maladies auto-immunes systémiques.

First-line treatments of autoimmune systemic diseases (ARD) are based on the use of various types of immunosuppressive or immunomodulatory drugs, either alone or in association, according to standardized reference protocols. Prolonged use of these drugs in severe or refractory ARD is associated with high morbidity and increased mortality. Innovative cell therapies represent a new promising approach for patients with ARDs, with the recent clinical use of: a) mesenchymal stromal cells (MSCs), based on their immunomodulatory, antifibrotic and pro-angiogenic properties and b) Chimeric Antigen Receptors (CAR) T cell therapies T lymphocytes, where genetically modified expression of a chimeric antigen receptor (CAR-T cells). Therapeutic use of MSC or CAR-T cells, remains indications of exception in patients with severe ARDs resistant to prior standard therapies with new prerequisite and organisation of health-care pathways as compared to traditional drugs, not only for the Cell and Gene Therapy (CGT) product definition and delivery process, but also for the patient clinical management before and after administration of the CGT product. The aim of this workshop under the auspices of the French Speaking Society of Bone Marrow and Cell transplantation (SFGM-TC) working group on autoimmune diseases (MATHEC) is to describe: a) the prerequisite for French hospitals to set-up the specific health-care pathways for MSC or CART therapy in ARDs patients, in accordance with regulatory and safety needs to perform academic or industry sponsored clinical trials, and b) the care-pathway for ARD patients treated with CGT, highlighting the importance of working in tandem between the ARD and the CAR-T cell specialist all along the indication, procedures and follow-up of ARDs. Patient safety considerations are central to guidance on patient selection to be validated collectively at the multidisciplinary team meeting (MDTM) based on recent (less than 3 months) thorough patient evaluation. MSC and CAR-T procedural aspects and follow-up are then carried out within appropriately experienced and SFGM-TC accredited centres in close collaboration with the ADs specialist.

Effectiveness of autologous emulsified stromal vascular fraction tissue injection for the treatment of complex perianal fistulas in inflammatory bowel diseases patients: a pilot study.

Complex fistulizing perianal disease is a disabling manifestation of inflammatory bowel disease (IBD), seriously compromising patients 'quality of life'. The success rate of available treatments is quite low, and nearly half of the patients will develop chronically active fistulas or experience fistula recurrence. Mesenchymal stem cell therapy has shown interesting results, but the complexity and the cost of production limit its widespread use. This study aims to report the results of the innovative use of autologous emulsified adipose-derived stromal vascular fraction tissue for treating complex fistulizing perianal disease. From March 2021 to March 2022, 10 patients underwent a two-step procedure: (1) examination under anaesthesia, with loose seton drainage and 4 weeks later and (2) curettage of the fistulous tract, internal fistula closure and an injection of autologous emulsified adipose-derived stromal vascular fraction tissue harvested from the subcutaneous layer of the patient's hip. Clinical and radiological (through magnetic resonance imaging) healing were assessed at 6 months. We included five patients affected by Crohn's disease, three by ulcerative colitis and two by indeterminate colitis. All patients were on concomitant biological therapy (50% on Infliximab). One patient required a re-treatment for a relapse and two different fistulas were separately treated in another one. Out of 12 total procedures performed, clinical healing was achieved in 10 cases (83%), while radiological healing in 6 patients (50%). No adverse events were recorded. Autologous emulsified adipose-derived stromal vascular fraction tissue can represent an effective, safe and cheap add-on therapy for patients with complex perianal fistulas in IBDs.

Engraftment of human mesenchymal stem cells in a severely immunodeficient mouse.

The transplantation of human mesenchymal stromal/stem cells (hMSCs) has potential as a curative and permanent therapy for congenital skeletal diseases. However, the self-renewal and differentiation capacities of hMSCs markedly vary. Therefore, cell proliferation and trilineage differentiation capacities were tested in vitro to characterize hMSCs before their clinical use. However, it remains unclear whether the ability of hMSCs in vitro accurately predicts that in living animals. The xenograft model is an alternative method for validating clinical MSCs. Nevertheless, the protocol still needs refinement, and it has yet to be established whether hMSCs, which are expanded in culture for clinical use, retain the ability to engraft and differentiate into adipogenic, osteogenic, and chondrogenic lineage cells in transplantation settings. In the present study, to establish a robust xenograft model of MSCs, we examined the delivery routes of hMSCs and the immunological state of recipients. The intra-arterial injection of hMSCs into X-ray-irradiated (IR) NOG, a severely immunodeficient mouse, achieved the highest engraftment but failed to sustain long-term engraftment. We demonstrated that graft cells localized to a collagenase-released fraction (CR), in which endogenous colony-forming cells reside. We also showed that Pdgfrα+Sca1+ MSCs (PαS), which reside in the CR fraction, resisted IR. These results show that our protocol enables hMSCs to fulfill a high level of engraftment in mouse bone marrow in the short term. In contrast, long-term reconstitution was restricted, at least partially, because of IR-resistant endogenous MSCs.

Transplantation of autologous mesenchymal stromal cells in complete cervical spinal cord injury: a pilot study.

Objective: Spinal cord injury (SCI) is a serious condition that can lead to partial or complete paraplegia or tetraplegia. Currently, there are few therapeutic options for these conditions, which are mainly directed toward the acute phase, such as surgical intervention and high-dose steroid administration. Mesenchymal stromal cells (MSC) have been shown to improve neurological function following spinal cord injury. The aim of the study was to evaluate the safety, feasibility, and potential efficacy of MSC transplantation in patients with cervical traumatic SCI. Methods: We included seven subjects with chronic traumatic SCI (> 1 year) at the cervical level, classified as American Spinal Cord Injury Association impairment scale (AIS) grade A. Subjects received two doses of autologous bone marrow derived MSC, the first by direct injection into the lesion site after hemilaminectomy and the second three months later by intrathecal injection. Neurologic evaluation, spinal magnetic resonance imaging (MRI), urodynamics, and life quality questionnaires were assessed before and after treatment. Results: Cell transplantation was safe without severe or moderate adverse effects, and the procedures were well tolerated. Neurological evaluation revealed discrete improvements in sensitivity below the lesion level, following treatment. Five subjects showed some degree of bilateral sensory improvement for both superficial and deep mechanical stimuli compared to the pretreatment profile. No significant alterations in bladder function were observed during this study. Conclusion: Transplantation of autologous MSC in patients with chronic cervical SCI is a safe and feasible procedure. Further studies are required to confirm the efficacy of this therapeutic approach. Clinical trial registration: https://clinicaltrials.gov/study/NCT02574572, identifier NCT02574572.

Heterogeneity in Dental Tissue-Derived MSCs Revealed by Single-Cell RNA-seq.

Mesenchymal stromal cells (MSCs) are multipotent, progenitor cells that reside in tissues across the human body, including the periodontal ligament (PDL) and gingiva. They are a promising therapeutic tool for various degenerative and inflammatory diseases. However, different heterogeneity levels caused by tissue-to-tissue and donor-to-donor variability, and even intercellular differences within a given MSCs population, restrict their therapeutic potential. There are considerable efforts to decipher these heterogeneity levels using different "omics" approaches, including single-cell transcriptomics. Previous studies applied this approach to compare MSCs isolated from various tissues of different individuals, but distinguishing between donor-to-donor and tissue-to-tissue variability is still challenging. In this study, MSCs were isolated from the PDL and gingiva of 5 periodontally healthy individuals and cultured in vitro. A total of 3,844 transcriptomes were generated using single-cell mRNA sequencing. Clustering across the 2 different tissues per donor identified PDL- and gingiva-specific and tissue-spanning MSCs subpopulations with unique upregulated gene sets. Gene/pathway enrichment and protein-protein interaction (PPI) network analysis revealed differences restricted to several cellular processes between tissue-specific subpopulations, indicating a limited tissue-of-origin variability in MSCs. Gene expression, pathway enrichment, and PPI network analysis across all donors' PDL- or gingiva-specific subpopulations showed significant but limited donor-to-donor differences. In conclusion, this study demonstrates tissue- and donor-specific variabilities in the transcriptome level of PDL- and gingiva-derived MSCs, which seem restricted to specific cellular processes. Identifying tissue-specific and tissue-spanning subpopulations highlights the intercellular differences in dental tissue-derived MSCs. It could be reasonable to control MSCs at a single-cell level to ensure their properties before transplantation.

Cytokine priming enhances the antifibrotic effects of human adipose derived mesenchymal stromal cells conditioned medium.

BACKGROUND: Fibrosis is a pathological scarring process characterized by persistent myofibroblast activation with excessive accumulation of extracellular matrix (ECM). Fibrotic disorders represent an increasing burden of disease-associated morbidity and mortality worldwide for which there are limited therapeutic options. Reversing fibrosis requires the elimination of myofibroblasts, remodeling of the ECM, and regeneration of functional tissue. Multipotent mesenchymal stromal cells (MSC) have antifibrotic properties mediated by secreted factors present in their conditioned medium (MSC-CM). However, there are no standardized in vitro assays to predict the antifibrotic effects of human MSC. As a result, we lack evidence on the effect of cytokine priming on MSC's antifibrotic effects. We hypothesize that the MSC-CM promotes fibrosis resolution in vitro and that this effect is enhanced following MSC cytokine priming. METHODS: We compared the antifibrotic effects of resting versus interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) primed MSC-CM in four in vitro assays: prevention of fibroblast activation, myofibroblasts deactivation, ECM degradation and fibrosis resolution in lung explant cultures. Furthermore, we performed transcriptomic analysis of myofibroblasts treated or not with resting or primed MSC-CM and proteomic characterization of resting and primed MSC-CM. RESULTS: We isolated MSC from adipose tissue of 8 donors, generated MSC-CM and tested each MSC-CM independently. We report that MSC-CM treatment prevented TGF-ß induced fibroblast activation to a similar extent as nintedanib but, in contrast to nintedanib, MSC-CM reduced fibrogenic myofibroblasts (i.e. transcriptomic upregulation of apoptosis, senescence, and inflammatory pathways). These effects were larger when primed rather than resting MSC-CM were used. Priming increased the ability of MSC-CM to remodel the ECM, reducing its content of collagen I and fibronectin, and reduced the fibrotic load in TGF-ß treated lung explant cultures. Priming increased the following antifibrotic proteins in MSC-CM: DKK1, MMP-1, MMP-3, follistatin and cathepsin S. Inhibition of DKK1 reduced the antifibrotic effects of MSC-CM. CONCLUSIONS: In vitro, MSC-CM promote fibrosis resolution, an effect enhanced following MSC cytokine priming. Specifically, MSC-CM reduces fibrogenic myofibroblasts through apoptosis, senescence, and by enhancing ECM degradation. Future studies will establish the in vivo relevance of MSC priming to fibrosis resolution.

A mathematical insight to control the disease psoriasis using mesenchymal stem cell transplantation with a biologic inhibitor.

Psoriasis is a chronic, non-contagious, immune-mediated skin disorder. Inflammation of the skin's surface is characterised by scaly white, red, or silvery spots that occur due to the hyper-proliferation of keratinocytes in the epidermal layer. Primarily, pharmaceutical drugs or immune therapy are used to treat psoriasis. We are all aware that, certain therapeutic strategies can have some adverse effects, and over time, that hidden inflammation may manifest. This article introduces a mathematical model for psoriasis, formulated by employing a set of nonlinear ordinary differential equations (ODEs) that describe the densities of T-cells, dendritic cells (DCs), keratinocytes, and mesenchymal stromal cells (MSCs) as basic cell populations. A tumor necrosis factor- α ( T N F - α ) inhibitor has been imposed from the initial stage of the treatment regime, using the optimal control theoretic approach, and the numerical results have been observed. After 80 days of monitoring using only biologic T N F - α inhibitors, if this approach did not provide the intended outcomes (when severity arises), stem cells are administered a few times in a pulsed manner as a cell replacement technique in addition to this anti T N F - α medicine. We have observed the combined therapeutic benefit of stem cell replacement with a T N F - α inhibitor from a mathematical point of view. The theoretical analysis and the numerical results revealed that stem cell transplantation, along with a T N F - α inhibitor, is a promising psoriasis treatment option moving forward.

Effects of Replicative Senescence of Human Chorionic MSCs on their EV-miRNA Profile.

Chorionic mesenchymal stromal cells (CHO-MSCs) and their extracellular vesicles (EVs) are becoming increasingly popular, since chorion is ethically harmless and an easily accessible source of MSCs. However, until now there is only a limited number of studies with a thorough characterization of CHO-MSCs derived EVs and their miRNA profile. In this study, we monitored changes in the EV-miRNA profile between early and late passage of human CHO-MSCs. First, senescence of CHO-MSCs was induced by serial passaging and confirmed by morphological changes, shortened telomeres and changes in the expression of selected genes. The expression of MSCs-specific surface markers CD73, CD90, CD105 did not change with increasing passages. Next, EVs and their miRNA profiles were compared between early vs late passage cells. Number of EVs and their size were not significantly changed. Seven of the top 10 most expressed EV-miRNAs were common to both early and late passages. A differential expression study between early and late passages identified 37 significantly differentially expressed EV-miRNAs, out of which 23 were found to be associated with pathways of cellular senescence based on KEGG pathway analysis. A set of 9 miRNAs were identified as the most frequently associated with senescence and/or with the most altered expression between early and late passages, out of which miR-145-5p, miR-335-5p and miR-199b-3p were the most significant downregulated miRNAs in late passages. The most upregulated EV-miRNAs were miR-1307-3p, miR-3615 and miR320b. Targeting these miRNAs in future experiments may prolong the therapeutic potential of CHO-MSCs and their EVs.