Amount Of Mesenchymal Stem Cells Research Articles

Young small extracellular vesicles rejuvenate replicative senescence by remodeling Drp1 translocation-mediated mitochondrial dynamics

BackgroundHuman mesenchymal stem cells have attracted interest in regenerative medicine and are being tested in many clinical trials. In vitro expansion is necessary to provide clinical-grade quantities of mesenchymal stem cells; however, it has been reported to cause replicative senescence and undefined dysfunction in mesenchymal stem cells. Quality control assessments of in vitro expansion have rarely been addressed in ongoing trials. Young small extracellular vesicles from the remnant pulp of human exfoliated deciduous teeth stem cells have demonstrated therapeutic potential for diverse diseases. However, it is still unclear whether young small extracellular vesicles can reverse senescence-related declines.ResultsWe demonstrated that mitochondrial structural disruption precedes cellular dysfunction during bone marrow-derived mesenchymal stem cell replication, indicating mitochondrial parameters as quality assessment indicators of mesenchymal stem cells. Dynamin-related protein 1-mediated mitochondrial dynamism is an upstream regulator of replicative senescence-induced dysfunction in bone marrow-derived mesenchymal stem cells. We observed that the application of young small extracellular vesicles could rescue the pluripotency dissolution, immunoregulatory capacities, and therapeutic effects of replicative senescent bone marrow-derived mesenchymal stem cells. Mechanistically, young small extracellular vesicles could promote Dynamin-related protein 1 translocation from the cytoplasm to the mitochondria and remodel mitochondrial disruption during replication history.ConclusionsOur findings show that Dynamin-related protein 1-mediated mitochondrial disruption is associated with the replication history of bone marrow-derived mesenchymal stem cells. Young small extracellular vesicles from human exfoliated deciduous teeth stem cells alleviate replicative senescence by promoting Dynamin-related protein 1 translocation onto the mitochondria, providing evidence for a potential rejuvenation strategy.Graphical

Extracellular vesicles derived from mesenchymal stem cells ameliorate cognitive impairment caused by neuroinflammation in young but not aged mice

Aim: The aim of this work was to study the effects of extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) on inflammation-impaired cognitive functions and the brain of mice. Methods: Young mice (~3-month-old) and aged mice (~18-month-old) were injected with bacterial lipopolysaccharide (LPS) and obtained intravenously donor 106 human umbilical cord MSCs, EVs isolated from a similar amount of MSCs or conditioned medium (CM) of MSCs. Subsequently, the mice were examined in behavioral tests and the mouse brains were analyzed for the levels of pro-inflammatory cytokines, α7 nicotinic acetylcholine receptors (α7 nAChRs) and amyloid beta 1-42 (Aβ1-42). Results: EVs prevented LPS-induced memory impairment in mice, whereas CM provided a weaker and temporal effect. Both EVs and MSCs injected once after regular injections of LPS stably improved memory of young mice. In contrast, both cells and EVs provided only transient effect in aged mice injected with LPS. The brains of aged LPS-treated mice contained elevated amounts of IL-1β and IL-6; both MSCs and EVs decreased them significantly. The brains of non-treated aged mice contained decreased levels of α7 nAChRs and increased levels of Aβ1-42 and α7-bound Aβ1-42 compared to the brains of young mice. LPS treatment decreased α7 nAChRs in both young and aged mice, while both MSCs and EVs restored them up to the control level. In young mice, LPS treatment increased the level of Aβ1-42 and α7-bound Aβ1-42, whereas MSCs and EVs decreased it. In contrast, neither LPS nor MSCs/EVs influenced the elevated level of Aβ1-42 but increased α7-bound Aβ1-42 in the brains of aged mice. Conclusions: Regenerative potential of MSCs and MSC-derived EVs is sufficient to support cognitive functions of LPS-treated young mice but is quite poor for aged animals, possibly, due to decreased levels of α7 nAChRs and accumulated Aβ1-42 in their brains.

Mesenchymal stem cells from bone marrow of patients with drug-resistant tuberculosis for cellular therapy

In this study, we evaluated the proliferative capacity of mesenchymal stem cells (MSCs) from the bone marrow of patients with drug-resistant tuberculosis (DR-TB) and selected the optimal conditions for obtaining autologous MSCs transplant. The results of the expansion of MSCs from 42 patients with DR-TB in medium with 10% FBS demonstrated low proliferative potential for of these cells compared with MSCs from 47 donors. MSCs content in 1 ml of bone marrow of patients was also lower. A relationship was found between the amount of MSCs in the primary culture and in the finished product, and with a small number of cells in the primary culture, we used 5% platelet lysate instead of 10% FTS, which made it possible to obtain a high dose of MSCs in the transplant.

Optimizing culture conditions of human umbilical cord mesenchymal stem cells (hUC-MSCs) by adjusting the volume of xeno-free supplement serum

Mesenchymal stem cells (MSCs) have become an effective tool for treating immune-related diseases due to their multilineage potential and immunomodulatory capabilities. However, a high cell dose is frequently essential for stem cell infusion in clinical practice. Therefore, it is necessary to produce sufficient quantities of MSCs while ensuring cell quality for clinical application in humans. To be able to use stem cells in patients requires a more rigorous captive procedure than using a xeno-free medium that does not contain substances derived from the hypothetical allergenic regime. Therefore, current cell culture procedures substitute xeno-free culture media with added supplement serum for the traditional DMEM media with bovine fetal serum (FBS). This switch increased the production cost immensely and made it difficult to produce MSCs on an industrial scale. In this study, we optimized the condition of MSCs’ cultures by adjusting the amount of the supplement serum usage to reduce production costs for industrial manufacturing. This is the first study to claim that reducing the amount of xeno-free supplement serum had no effect on the quality of hUC-MSCs isolated from Vietnamese children’s umbilical cords.

Large-scale expansion of human umbilical cord-derived mesenchymal stem cells using PLGA@PLL scaffold

Mesenchymal stem cells (MSCs) are highly important in biomedicine and hold great potential in clinical treatment for various diseases. In recent years, the capabilities of MSCs have been under extensive investigation for practical application. Regarding therapy, the efficacy usually depends on the amount of MSCs. Nevertheless, the yield of MSCs is still limited due to the traditional cultural methods. Herein, we proposed a three-dimensional (3D) scaffold prepared using poly lactic-co-glycolic acid (PLGA) nanofiber with polylysine (PLL) grafting, to promote the growth and proliferation of MSCs derived from the human umbilical cord (hUC-MSCs). We found that the inoculated hUC-MSCs adhered efficiently to the PLGA scaffold with good affinity, fast growth rate, and good multipotency. The harvested cells were ideally distributed on the scaffold and we were able to gain a larger yield than the traditional culturing methods under the same condition. Thus, our cell seeding with a 3D scaffold could serve as a promising strategy for cell proliferation in the large-scale production of MSCs. Moreover, the simplicity and low preparation cost allow this 3D scaffold to extend its potential application beyond cell culture.Graphical

Efecto de la diabetes en células madres de la cavidad bucal. Una revisión breve

Tissue engineering is one of the pillars of regeneration techniques in medicine, based on the use of mesenchymal stem cells (MSCs). MSCs have self-renewal and differentiation capabilities in multiple lines, due to which they are multipotent mature cells. MSCs can be spread over several tiles, a good amount of MSCs is in the oral cavity, including parts such as the alveolar bone, periodontal ligament, dental pulp, dental follicle, oral mucosa and gums. Despite being a major advance in medical sciences, the use of MSCs may be limited by several factors that affect the biological capabilities of these cells. Considering the oral cavity as one of the main sources of obtaining a high amount MSCs, one of the factors that has great repercussions on oral health and diabetes, the clearest example is the two-way relationship that has periodontitis today. periodontitis is considered the sixth complication of diabetes. Given the above, it is necessary to identify the effects thatcould also have on the MSCs obtained in the oral cavity, effects that could hinder their use in tissue engineering. The purpose of this review is to describe the effect of diabetes on MSCs in the oral cavity.

In vivo non-invasive monitoring of tissue development in 3D printed subcutaneous bone scaffolds using fibre-optic Raman spectroscopy

The development of novel biomaterials for regenerative therapy relies on the ability to assess tissue development, quality, and similarity with native tissue types in in vivo experiments. Non-invasive imaging modalities such as X-ray computed tomography offer high spatial resolution but limited biochemical information while histology and biochemical assays are destructive. Raman spectroscopy is a non-invasive, label-free and non-destructive technique widely applied for biochemical characterization. Here we demonstrate the use of fibre-optic Raman spectroscopy for in vivo quantitative monitoring of tissue development in subcutaneous calcium phosphate scaffolds in mice over 16 weeks. Raman spectroscopy was able to quantify the time dependency of different tissue components related to the presence, absence, and quantity of mesenchymal stem cells. Scaffolds seeded with stem cells produced 3–5 times higher amount of collagen-rich extracellular matrix after 16 weeks implantation compared to scaffolds without. These however, showed a 2.5 times higher amount of lipid-rich tissue compared to implants with stem cells. Ex vivo micro-computed tomography and histology showed stem cell mediated collagen and bone development. Histological measures of collagen correlated well with Raman derived quantifications (correlation coefficient in vivo 0.74, ex vivo 0.93). In the absence of stem cells, the scaffolds were largely occupied by adipocytes. The technique developed here could potentially be adapted for a range of small animal experiments for assessing tissue engineering strategies at the biochemical level.

Different Sources of Mesenchymal Stem Cells for Tissue Regeneration: A Guide to Identifying the Most Favorable One in Orthopedics and Dentistry Applications.

The success of regenerative medicine in various clinical applications depends on the appropriate selection of the source of mesenchymal stem cells (MSCs). Indeed, the source conditions, the quality and quantity of MSCs, have an influence on the growth factors, cytokines, extracellular vesicles, and secrete bioactive factors of the regenerative milieu, thus influencing the clinical result. Thus, optimal source selection should harmonize this complex setting and ensure a well-personalized and effective treatment. Mesenchymal stem cells (MSCs) can be obtained from several sources, including bone marrow and adipose tissue, already used in orthopedic regenerative applications. In this sense, for bone, dental, and oral injuries, MSCs could provide an innovative and effective therapy. The present review aims to compare the properties (proliferation, migration, clonogenicity, angiogenic capacity, differentiation potential, and secretome) of MSCs derived from bone marrow, adipose tissue, and dental tissue to enable clinicians to select the best source of MSCs for their clinical application in bone and oral tissue regeneration to delineate new translational perspectives. A review of the literature was conducted using the search engines Web of Science, Pubmed, Scopus, and Google Scholar. An analysis of different publications showed that all sources compared (bone marrow mesenchymal stem cells (BM-MSCs), adipose tissue mesenchymal stem cells (AT-MSCs), and dental tissue mesenchymal stem cells (DT-MSCs)) are good options to promote proper migration and angiogenesis, and they turn out to be useful for gingival, dental pulp, bone, and periodontal regeneration. In particular, DT-MSCs have better proliferation rates and AT and G-MSC sources showed higher clonogenicity. MSCs from bone marrow, widely used in orthopedic regenerative medicine, are preferable for their differentiation ability. Considering all the properties among sources, BM-MSCs, AT-MSCs, and DT-MSCs present as potential candidates for oral and dental regeneration.

Mesenchymal stem cells protect against acetaminophen hepatotoxicity by secreting regenerative cytokine hepatocyte growth factor

BackgroundAcetaminophen (APAP) overdose is a major cause of the morbidity of acute liver failure. The current clinically approved treatment for APAP poisoning, N-acetylcysteine (NAC), has a limited therapeutic window, and prolonged treatment with NAC delays liver regeneration. Mesenchymal stem cells (MSCs) also have therapeutic effects on APAP-induced mouse liver failure, but whether the effects are comparable to those of NAC has not been determined, and the mechanism still needs further exploration.MethodsFasted C57BL/6 mice that received 500 mg/kg APAP were treated intravenously with 300 mg/kg NAC or different amounts of MSCs at 2 h after APAP to investigate survival, hepatocyte necrosis and neutrophil/macrophage recruitment. In vitro co-culture was performed to study the anti-necrotic effects of MSCs on the APAP-injured hepatocyte cell line L-O2.ResultsMSCs dose-dependently rescued the C57BL/6J mice from APAP-induced liver failure, with 87.5% of MSCs (1 × 106) surviving similar to that of NAC (90%). MSC has similar effects on reduced hepatocyte necrosis and granulocytic myeloid-derived suppressor cells (MDSC) infiltration but enhanced the proportion of regenerative monocytic MDSC when compared to NAC. Mechanistically, MSCs attenuate hepatocyte necrosis by secreting hepatocyte growth factor (HGF). When HGF was knocked down, the protective effects of MSCs were reduced on APAP-induced hepatocyte necrosis and mouse liver failure.ConclusionsMSCs are comparable to NAC against APAP-induced liver failure by secreting HGF with less regenerative retardation concerns, thus facilitating the application of MSCs in clinical therapy for APAP liver failure.

Subconjunctival use of allogeneic mesenchymal stem cells to treat chronic superficial keratitis in German shepherd dogs: Pilot study.

Chronic superficial keratitis (CSK) is an ocular condition in dogs characterized by corneal opacification leading to visual function impairment. Control of this chronic condition requires the use of topical immunomodulators or corticosteroids daily. Regenerative medicine has shown promising results in several fields of medicine. The aim of this study was to evaluate the clinical effect of allogeneic mesenchymal stem cells (MSCs) of adipose tissue applied via subconjunctival in dogs with CSK. A series of cases of eight dogs diagnosed with CSK were divided into two groups, four dogs each; the conventional treatment group received prednisolone 1% as topical eye drops and the experimental group (EG) received allogeneic MSCs transplantation. The dogs had not previously been treated for CSK. Systemic and ophthalmologic examinations were performed to exclude other abnormalities. An administered amount of MSC (1 × 106 cells each time) was injected via subconjunctival in the peri-limbal region at 0 and 30 days. The animals were followed for 110 days for clinical evaluation, and, at the same time, the images of the corneal abnormalities were obtained and analyzed in the ImageJ software. The statistical analysis was performed in the GrandPrism 7.0 software. Initial and final images revealed that areas with neovascularization, inflammatory infiltrate, and opacity regressed in most eyes in both groups (7/8 eyes in each group) at the end of the 110 days, p = 0.0391 and p = 0.0078 respectively, but this response was minor in the EG comparing to conventional group (CG) (p = 0.026). No local or systemic side effects were observed. Despite the small melioration, MSCs treatment suggests clinical improvement in patients with CSK after 110 days without any local or systemic side effects. However, the improvement achieved was significantly less than the observed within CG. Further studies still are needed to evaluate the use and benefits of stem cells as an adjunct treatment for CSK.

Human ESC-derived Neuromesodermal Progenitors (NMPs) Successfully Differentiate into Mesenchymal Stem Cells (MSCs).

Mesenchymal Stem Cells (MSCs), as an adult stem cell type, are used to treat various disorders in clinics. However, derivation of homogenous and adequate amount of MSCs limits the regenerative treatment potential. Although mesoderm is the main source of mesenchymal progenitors during embryonic development, neuromesodermal progenitors (NMPs), reside in the primitive streak during development, is known to differentiate into paraxial mesoderm. In the current study, we generated NMPs from human embryonic stem cells (hESC), subsequently derived MSCs and characterized this cell population in vitro and in vivo. Using a bFGF and CHIR induced NMP formation protocol followed by serum containing culture conditions; here we show that MSCs can be generated from NMPs identified by not only the expression of T/Bra and Sox 2 but also FLK-1/PDGFRα in our study. NMP-derived MSCs were plastic adherent fibroblast like cells with colony forming capacity and trilineage (osteo-, chondro- and adipo-genic) differentiation potential. In the present study, we demonstrate that NMP-derived MSCs have an endothelial tendency which might be related to their FLK-1+/PDGFRα + NMP origin. NMP-derived MSCs displayed a protein expression profile ofcharacterized MSCs. Growth factor and angiogenesis related pathway proteins were similarly expressed in NMP-derived MSCs and characterized MSCs. NMP-derived MSCs keep characteristics after short-term and long-term freeze-thaw cycles and localized into bone marrow followed by tail vein injection into NOD/SCID mice. Together, these data showed that hESC-derived NMPs might be used as a precursor cell population for MSC derivation and could be used for in vitro and in vivo research.

Enhancement of bone marrow aspirate concentrate with local self-healing corticotomies

Bone marrow aspirate concentrate (BMAC) is a potentially useful biological product for bone regeneration. This study investigated whether BMAC can be enriched by local minor corticotomies. Five 4-month-old domestic pigs were used with each pig undergoing two minor corticotomies at one randomly-selected tibia. Two weeks after the operation, bone marrow was aspirated from both tibiae and processed into BMAC samples. The amount of mesenchymal stem cells (MSCs) and the concentration of several regenerative growth factors contained in BMAC, as well as the proliferative and osteogenic differentiation capacity of MSCs, were compared between the corticotomy and the control sides. Another four weeks later, healing of the corticotomies was evaluated by radiographic and histological methods. The results demonstrated that BMAC from the corticotomy side contained significantly more MSCs than the control side. MSCs from the corticotomy side also proliferated significantly faster and tended to have stronger osteogenic differentiation than those from the control side. In contrast, the protein concentration of TGF-β, BMP-2 and PDGF contained in BMAC was only minimally changed by the corticotomies. The corticotomies in all pigs healed uneventfully, showing complete obliteration of the corticotomy gaps on CT images. Comparison between the two sides showed that the corticotomy side had thicker and denser cortical bone and more abundant osteogenic cell differentiation than the control side. These findings suggest that the quantity and proliferative/osteogenic differentiation capacity of MSCs contained in local BMAC can be enhanced by minor corticotomies, and spontaneous healing of the corticotomy can be completed within 6 weeks of the operation.

Melatonin suppresses senescence-derived mitochondrial dysfunction in mesenchymal stem cells via the HSPA1L-mitophagy pathway.

Mesenchymal stem cells (MSCs) are a popular cell source for stem cell‐based therapy. However, continuous ex vivo expansion to acquire large amounts of MSCs for clinical study induces replicative senescence, causing decreased therapeutic efficacy in MSCs. To address this issue, we investigated the effect of melatonin on replicative senescence in MSCs. In senescent MSCs (late passage), replicative senescence decreased mitophagy by inhibiting mitofission, resulting in the augmentation of mitochondrial dysfunction. Treatment with melatonin rescued replicative senescence by enhancing mitophagy and mitochondrial function through upregulation of heat shock 70 kDa protein 1L (HSPA1L). More specifically, we found that melatonin‐induced HSPA1L binds to cellular prion protein (PrPC), resulting in the recruitment of PrPC into the mitochondria. The HSPA1L‐PrPC complex then binds to COX4IA, which is a mitochondrial complex IV protein, leading to an increase in mitochondrial membrane potential and anti‐oxidant enzyme activity. These protective effects were blocked by knockdown of HSPA1L. In a murine hindlimb ischemia model, melatonin‐treated senescent MSCs enhanced functional recovery by increasing blood flow perfusion, limb salvage, and neovascularization. This study, for the first time, suggests that melatonin protects MSCs against replicative senescence during ex vivo expansion for clinical application via mitochondrial quality control.

Full title: High glucose protects mesenchymal stem cells from metformin-induced apoptosis through the AMPK-mediated mTOR pathway

Micro- and macro-vascular events are directly associated with hyperglycemia in patients with type 2 diabetes mellitus (T2DM), but whether intensive glucose control decreases the risk of diabetic cardiovascular complications remains uncertain. Many studies have confirmed that impaired quality and quantity of mesenchymal stem cells (MSCs) plays a pathogenic role in diabetes. Our previous study found that the abundance of circulating MSCs was significantly decreased in patients with T2DM, which was correlated with the progression of diabetic complications. In addition, metformin-induced MSC apoptosis is one of the reasons for the decreased quantity of endogenous or exogenous MSCs during intensive glucose control. However, the role of glucose in metformin-induced MSC apoptosis during intensive glucose control in T2DM remains unknown. In this study, we found that metformin induces MSC apoptosis during intensive glucose control, while high glucose (standard glucose control) could significantly reverse its adverse effect in an AMPK-mTOR pathway dependent manner. Thus, our results indicate that the poorer clinical benefit of the intensive glucose control strategy may be related to an adverse effect due to metformin-induced MSC apoptosis during intensive glucose control therapy in patients with T2DM.

Comparable effect of adipose-derived stromal vascular fraction and mesenchymal stem cells for wound healing: An in vivo study

Introduction: Mesenchymal stem cells (MSCs) have been known to have angiogenic potency, particularly for wound healing. However, it is such a cumbersome procedure to obtain a large amount of MSCs due to the culture process. The stromal vascular fraction (SVF), or so called nonexpanded MSCs, of adipose tissue is gaining increased interest in the field of cell therapy. An optimized method that takes only 2 hours to produce a large amount of stromal cells from adipose tissue has been developed. Burn wound model was used to assess the efficacy of SVF produced by our method. This study aimed to evaluate the efficacy of SVF compared to MSCs. Methods: Male Sprague-Dawley rats with second degree burn wound were divided into the study groups: SVF, MSCs and saline control. The wound was photographed and evaluated weekly up to 5 weeks. The comparative analysis consisted of healing time, histology of skin tissue, as well as rat and human vascular endothelial growth factor (VEGF) expression. Results: In this study, wounds treated with SVF and MSCs were smaller after 14-21 days, compared to the untreated group. Expression of rat VEGF in SVF and MSC- treated groups after seven days post-wounded were also higher than the untreated group. No human VEGF was found expressed in the skin biopsies. Conclusion: This suggests that SVF and MSCs promote wound healing via a paracrine effect. The results suggest that SVF may be useful for wound healing and may be used as a promising alternative to MSC-based therapy.  

Evaluation of alginate hydrogel encapsulated mesenchymal stem cell migration in horses

Osteoarthritis is an incapacitating disease characterized by pain and a progressive decrease in joint mobility. The implantation of mesenchymal stem cells (MSCs) has shown promising results for its treatment. The challenge remains to keep the cells longer at the site of action, increasing their therapeutic potential. The aim of this study was to evaluate the effectiveness of the Qtracker® 655 nanocrystal marking on allogeneic synovial membrane (SM) MSCs, encapsulated in alginate hydrogel, evaluating the migration of these cells. The 10 radiocarpal joints were submitted to arthroscopic surgery (D0), divided into two groups. The chondral defect was treated according to the group: GA free-labelled MSCSM and GB labelled MSCSM microcapsules. Seven days after lesion induction and implantation of labelled cells, biopsies of the lesion site were performed in two animals, and fragments of SM and joint capsule also collected, which were frozen and later processed for fluorescence microscopy. The synovial fluid of the three animals was analyzed by flow cytometry three times – 3, 7 and 21 days after application. The cellular marking with the nanocrystals allowed the visualization of the cells in cartilage, synovial membrane, synovial fluid and articular capsule, but with a predilection for the synovial membrane and the lesion site was scarce. The labelled MSCSM in microcapsules were scarce in the synovial fluid and could be related to the small quantity of MSCs leaving the pores of the microcapsules, also favorable results, as the cells release paracrine effects acting for a long period until the cellular differentiation.

Abstract WP141: Neuroprotection by Mesenchymal Stem Cells (MSC) Administration Was Enhanced by Local Cooling Infusion (LCI) in Ischemic Rats

Introduction: Although mitochondrial transfer from MSCs to neurons has been found to be a promising strategy for the treatment of ischemic stroke, uncertainty regarding the appropriate quantity of MSCs and inefficient delivery to affected tissue have limited the applicability of this emerging therapy. In this study we sought, using LCI-mediated hypothermia, to provide a micro-environment conducive to mitochondrial homing and function, and thus to improve the neuroprotective efficacy of MSC mitochondrial transfer. Methods: Sprague-Dawley rats were subjected to 90min middle cerebral artery occlusion followed by 24h reperfusion. Rats were treated with either MSCs (1х10 5 ), LCI (cold saline, 0.6 ml/min, 5min) or both. After 24h reperfusion, infarct volume and neurological deficits were examined. After 28d, long-term functional outcomes were evaluated using foot-fault and rota-rod testing. In parallel, human neural SH-SY5Y cells were investigated in vitro using 2h oxygen-glucose deprivation (OGD), followed by MSC co-culture, with or without hypothermia (34°C, 4h). Mitochondrial transfer was assessed by O 2 consumption, extracellular ATP levels, mitochondrial membrane potential (MMP) in co-culture supernatant, and quantity (Western blot) of mitochondrial transfer related protein Rho GTPase Miro 1. Neural cell viability and intracellular ATP were measured at 6 h and 24 h after OGD. Results: After 24h reperfusion, MSC and LCI groups showed significant decreases in infarct volume and neuronal deficit. After 28d, the combination therapy group showed significantly greater long-term recovery than either of the monotherapy groups. With hypothermia, cell culture showed increased extracellular ATP, MMP and O 2 consumption in supernatant, cell viability, and neuronal intracellular ATP, indicating that MSC mitochondrial transfer to neurons was enhanced by hypothermia. Miro 1 was observed to be upregulated in the combination group, indicating its involvement in the transfer mechanism. Conclusion: Therapeutic hypothermia, by upregulating Miro 1 and thus improving transfer, greatly enhanced MSC mitochondrial transfer-mediated neuroprotection in ischemic stroke. This combined approach may facilitate the clinical translation of this therapy.

Noninvasive application of mesenchymal stem cell spheres derived from hESC accelerates wound healing in a CXCL12-CXCR4 axis-dependent manner.

Mesenchymal stem cells (MSC) derived from adult tissues effectively promote wound healing. However, MSC quality varies, and the quantity of MSC is limited, as MSC are acquired through donations. Moreover, the survival and functioning of dissociated MSC delivered to an inflammatory lesion are subject to challenges.Methods: Here, spheres (EMSCSp) generated from human embryonic stem cell-derived MSC (EMSC) were directly dropped onto excised wounds in mice; the effects of EMSCSp were compared to those of dissociated EMSC (EMSCDiss). Following transplantation, we measured the extent of wound closure, dissected the histological features of the wounds, determined transcriptomic changes in cells isolated from the treated and control wounds, and evaluated the molecular mechanism of the effects of EMSC.Results: The application of EMSCSp onto murine dermal wounds substantially increased survival and efficacy of EMSC compared to the topical application of EMSCDiss. RNA sequencing (RNA-Seq) of cells isolated from the wounds highlighted the involvement of CXCL12-CXCR4 signaling in the effects of EMSCSp, which was verified in EMSC via CXCL12 knockdown and in target cells (vascular endothelial cells, epithelial keratinocytes, and macrophages) via CXCR4 inhibition. Finally, we enhanced the biosafety of EMSCSp by engineering cells with an inducible suicide gene.Conclusions: Together, these data suggest the topical application of EMSCSp as an unlimited, quality-assured, safe, and noninvasive therapy for wound healing and the CXCL12-CXCR4 axis as a key player in this treatment.

Isolation of Adipose Tissue-Derived Stem Cells: Enzymatic Digestion in Combination with Mechanical Distortion to Increase Adipose Tissue-Derived Stem Cell Yield from Human Aspirated Fat.

Mesenchymal stem cells (MSCs) are of great interest due to their properties of immune modulation, tissue regeneration, and multipotent differentiation. Future developments of clinical applications, however, require a higher yield of MSCs, lower number of passages of cells in culture, and shorter time from harvest to use. Optimization and standardization of techniques for mesenchymal adipose tissue-derived stem cell isolation offers solutions to current bottlenecks as a larger amount of MSCs can be isolated. These improvements result in shorter expansion time, fewer passages, less donor material needed, and higher MSC yield. This paper describes an MSC isolation method combining enzymatic digestion with mechanic disruption. This protocol is a standardized and easy-to-implement method for reaching significantly higher MSC yields compared to conventional enzymatic isolation protocols. Based on the results presented, we hypothesize that the combined enzymatic and mechanical method increases the surface area of the adipose tissue, facilitating digestion by enzymes. This approach reduces the amount of adipose tissue and in vitro expansion time needed to reach sufficient amounts of MSCs for clinical purposes. Importantly, the method does not require increased amounts of collagenase, nor does it impair the viability or differentiability of the MSCs. Using this protocol increases MSC yield by a factor of three. As a consequence, these results indicate that the physiological concentration of MSCs in adipose tissue is higher than previously assumed. © 2018 by John Wiley & Sons, Inc.

Characterization and cost-benefit analysis of automated bioreactor-expanded mesenchymal stem cells for clinical applications.

Expanding quantities of mesenchymal stem cells (MSCs) sufficient to treat large numbers of patients in cellular therapy and regenerative medicine clinical trials is an ongoing challenge for cell manufacturing facilities. We evaluated options for scaling up large quantities of bone marrow-derived MSCs (BM-MSCs) using methods that can be performed in compliance with Good Manufacturing Practices (GMP). We expanded BM-MSCs from fresh marrow aspirate in αMEM supplemented with 5% human platelet lysate using both an automated cell expansion system (Quantum, Terumo BCT) and a manual flask-based method using multilayer flasks. We compared MSCs expanded using both methods and assessed their differentiation to adipogenic and osteogenic tissue, capacity to suppress T-cell proliferation, cytokines, and growth factor secretion profile and cost-effectiveness of manufacturing enough BM-MSCs to administer a single dose of 100 × 106 cells per subject in a clinical trial of 100 subjects. We have established that large quantities of clinical-grade BM-MSCs manufactured with an automated hollow-fiber bioreactor were phenotypically (CD73, CD90, CD105) and functionally (adipogenic and osteogenic differentiation and cytokine and growth factor secretion) similar to manually expanded BM-MSCs. In addition, MSC manufacturing costs significantly less and required less time and effort when using the Quantum automated cell expansion system over the manual multilayer flasks method. MSCs manufactured by an automated bioreactor are physically and functionally equivalent to the MSCs manufactured by the manual flask method and have met the standards required for clinical application.