Exosomes derived from CD271+CD56+ bone marrow mesenchymal stem cell subpopoulation identified by single-cell RNA sequencing promote axon regeneration after spinal cord injury.

Rationale: Spinal cord injury (SCI) results in neural tissue damage. However, the limited regenerative capacity of adult mammals' axons upon SCI leads to persistent neurological dysfunction. Thus, exploring the pathways that can enhance axon regeneration in injured spinal cord is of great significance. Methods: Through the utilization of single-cell RNA sequencing in this research, a distinct subpopulation of bone marrow mesenchymal stem cells (BMSCs) that exhibits the capacity to facilitate axon regeneration has been discovered. Subsequently, the CD271+CD56+ BMSCs subpopulation was isolated using flow cytometry, and the exosomes derived from this subpopulation (CD271+CD56+ BMSC-Exos) were extracted and incorporated into a hydrogel to create a sustained release system. The aim was to investigate the therapeutic effects of CD271+CD56+ BMSC-Exos and elucidate the underlying mechanisms involved in promoting axon regeneration and neural function recovery. Results: The findings indicate that CD271+CD56+ BMSC-Exos share similar physical and chemical properties with conventional exosomes. Importantly, in an SCI model, in situ implantation of CD271+CD56+ BMSC-Exos hydrogel resulted in increased expression of NF and synaptophysin, markers associated with axon regeneration and synapse formation, respectively. This intervention also contributed to improved neural function recovery. In vitro experiments demonstrated that CD271+CD56+ BMSC-Exos treatment significantly enhanced axon extension distance and increased the number of branches in dorsal root ganglion axons. Moreover, further investigation into the molecular mechanisms underlying CD271+CD56+ BMSC-Exos-mediated axon regeneration revealed the crucial involvement of the miR-431-3p/RGMA axis. Conclusion: In summary, the implantation of CD271+CD56+ BMSC-Exos hydrogel presents a promising and effective therapeutic approach for SCI.

Delayed Wound Healing in the Elderly and a New Therapeutic Target: CD271.

With the development of society, the global population is showing a trend of aging. It is well known that age is one of the factors affecting wound healing. Aging compromises the normal physiological process of wound healing, such as the change of skin structure, the decrease of growth factors, the deceleration of cell proliferation, and the weakening of migration ability, hence delaying wound healing. At present, research in adult stem cell-related technology and its derived regenerative medicine provides a novel idea for the treatment of senile wounds. Studies have confirmed that CD271 (P75 neurotropism receptor/P75NTR)-positive cells (CD271+ cells) are a kind of stem cells with a stronger ability of proliferation, differentiation, migration and secretion than CD271 negative (CD271- cells). Meanwhile, the total amount and distribution of CD271 positive cells in different ages of skin are also different, which may be related to the delayed wound healing of aging skin. Therefore, this article reviews the relationship between CD271+ cells and senile wounds and discusses a new scheme for the treatment of senile wounds.

Expression profiles of human somatic mesenchymal stem cells derived from fresh endometrium, ectopic-endometrium and umbilical cord.

OBJECTIVES: The study investigated the stem cell expression profiles and differentiation capacities of mesenchymal stem cells (MSCs) from different tissues, specifically human eutopic endometrium MSCs (eut-MSCs), ectopic endometrium MSCs (ect-MSCs), and umbilical cord MSCs (UC-MSCs). Our aim was to identify any similarities in subpopulations among these MSCs and lay a foundation for MSCs repair. MATERIAL AND METHODS: MSCs were isolated from endometrial tissue (n = 5), endometriosis tissue (n = 6), and umbilical cords (n = 7). Flow cytometry was used to examine cell phenotype, and three lineage tests were conducted to evaluate the differentiation capacity of the MSCs. RESULTS: Eut-MSCs expressed CD44 (98.00 ± 0.96%), CD73 (99.54 ± 0.02%), CD140b (99.16 ± 0.50%), CD146 (93.87 ± 2.27%), SUSD2 (50.76 ± 8.15%), and CD271 (2.1 ± 1.22%). Ect-MSCs expressed CD44 (98.23 ± 1.60%), CD73 (99.63 ± 0.04%), CD140b (98.13 ± 0.53%), CD146 (93.88 ± 3.19%), SUSD2 (49.33 ± 6.36%), and CD271 (2.85 ± 1.17%). UC-MSCs expressed CD44 (99.11 ± ± 0.42%), CD73 (99.65 ± 0.12%), CD140b (99.84 ± 0.42%), CD146 (88.09 ± 4.20%), SUSD2 (72.87 ± 7.13%), and CD271 (6.19 ± 2.08%). The expression of SUSD2 and CD271 in UC-MSCs was slightly but not significantly higher than that in ect-MSCs and eut-MSCs. However, CD44, CD73, CD140b, and CD146 showed similar expression levels in UC-MSCs, ect-MSCs, and eut-MSCs. All three types of MSCs demonstrated the capacity to differentiate into osteoblasts, adipocytes, and chondrocytes. CONCLUSIONS: Our findings indicate that ect-MSCs, eut-MSCs, and UC-MSCs have similar stem cell phenotypes and the ability to differentiate into three lineages.

Putative Cancer Stem Cell Markers are Frequently Expressed by Melanoma Cells in Vitro and in Situ but are also Present in Benign Differentiated Cells.

BACKGROUND: Currently, there remains an incomplete view of cancer stem cells (CSCs) in solid tumours. METHODS: We studied a panel of putative CSC surface markers (ALDH1A1, ABCG2, CD44v7/8, CD44v10, CD133, CD271, and Nestin) in 40 established melanoma cell lines and four early-passage melanoma strains by flow cytometry. We additionally examined 40 formalin-fixed paraffin-embedded melanoma tissues using immunofluorescence microscopy. This was compared with their expression in healthy skin, normal differentiated melanocytes and fibroblasts. RESULTS: Most of the putative CSC markers were expressed by both melanoma cell lines and tissues. When present, these proteins were expressed by the majority of cells in the population. However, the expression of these markers by cells in healthy skin sections, normal differentiated melanocytes, and fibroblasts revealed that differentiated non-malignant cells also expressed CSC markers indicating that they lack of specificity for CSCs. Culturing cell lines under conditions more characteristic of the tumour microenvironment upregulated CSC marker expressions in a proportion of cell lines, which correlated with improved cell growth and viability. CONCLUSIONS: The testing of melanoma cell lines (n = 40), early-passage cell strains (n = 4), and melanoma tissues (n = 40) showed that several putative CSC markers (ALDH1A1, ABCG2, CD44v7/8, CD44v10, CD133, CD271, and Nestin) are commonly present in a large proportion of melanoma cells in vitro and in situ. Further, we showed that these putative markers lack specificity for CSCs because they are also expressed in differentiated non-malignant cell types (melanocytes, fibroblasts, and skin), which could limit their use as therapeutic targets. These data are consistent with the emerging notion of CSC plasticity and phenotype switching within cancer cell populations.

Synovial membrane-derived mesenchymal progenitor cells from osteoarthritic joints in dogs possess lower chondrogenic-, and higher osteogenic capacity compared to normal joints.

BACKGROUND: Synovial membrane-derived mesenchymal progenitor cells (SM-MPCs) are a promising candidate for the cell-based treatment of osteoarthritis (OA) considering their in vitro and in vivo capacity for cartilage repair. However, the OA environment may adversely impact their regenerative capacity. There are no studies for canine (c)SM-MPCs that compare normal to OA SM-MPCs, even though dogs are considered a relevant animal model for OA. Therefore, this study compared cSM-MPCs from normal and OA synovial membrane tissue to elucidate the effect of the OA environment on MPC numbers, indicated by CD marker profile and colony-forming unit (CFU) capacity, and the impact of the OA niche on tri-lineage differentiation. METHODS: Normal and OA synovial membrane were collected from the knee joints of healthy dogs and dogs with rupture of the cruciate ligaments. The synovium was assessed by histopathological OARSI scoring and by RT-qPCR for inflammation/synovitis-related markers. The presence of cSM-MPCs in the native tissue was further characterized with flow cytometry, RT-qPCR, and immunohistochemistry, using the MPC markers; CD90, CD73, CD44, CD271, and CD34. Furthermore, cells isolated upon enzymatic digestion were characterized by CFU capacity, and a population doublings assay. cSM-MPCs were selected based on plastic adherence, expanded to passage 2, and evaluated for the expression of MPC-related surface markers and tri-lineage differentiation capacity. RESULTS: Synovial tissue collected from the OA joints had a significantly higher OARSI score compared to normal joints, and significantly upregulated inflammation/synovitis markers S100A8/9, IL6, IL8, and CCL2. Both normal and OA synovial membrane contained cells displaying MPC properties, including a fibroblast-like morphology, CFU capacity, and maintained MPC marker expression over time during expansion. However, OA cSM-MPCs were unable to differentiate towards the chondrogenic lineage and had low adipogenic capacity in contrast to normal cSM-MPCs, whereas they possessed a higher osteogenic capacity. Furthermore, the OA synovial membrane contained significantly lower percentages of CD90+, CD44+, CD34+, and CD271+ cells. CONCLUSIONS: The OA environment had adverse effects on the regenerative potential of cSM-MPCs, corroborated by decreased CFU, population doubling, and chondrogenic capacity compared to normal cSM-MPCs. OA cSM-MPCs may be a less optimal candidate for the cell-based treatment of OA than normal cSM-MPCs.

Full spectrum flow cytometry reveals mesenchymal heterogeneity in first trimester placentae and phenotypic convergence in culture, providing insight into the origins of placental mesenchymal stromal cells.

Single-cell technologies (RNA-sequencing, flow cytometry) are critical tools to reveal how cell heterogeneity impacts developmental pathways. The placenta is a fetal exchange organ, containing a heterogeneous mix of mesenchymal cells (fibroblasts, myofibroblasts, perivascular, and progenitor cells). Placental mesenchymal stromal cells (pMSC) are also routinely isolated, for therapeutic and research purposes. However, our understanding of the diverse phenotypes of placental mesenchymal lineages, and their relationships remain unclear. We designed a 23-colour flow cytometry panel to assess mesenchymal heterogeneity in first-trimester human placentae. Four distinct mesenchymal subsets were identified; CD73+CD90+ mesenchymal cells, CD146+CD271+ perivascular cells, podoplanin+CD36+ stromal cells, and CD26+CD90+ myofibroblasts. CD73+CD90+ and podoplanin + CD36+ cells expressed markers consistent with cultured pMSCs, and were explored further. Despite their distinct ex-vivo phenotype, in culture CD73+CD90+ cells and podoplanin+CD36+ cells underwent phenotypic convergence, losing CD271 or CD36 expression respectively, and homogenously exhibiting a basic MSC phenotype (CD73+CD90+CD31-CD144-CD45-). However, some markers (CD26, CD146) were not impacted, or differentially impacted by culture in different populations. Comparisons of cultured phenotypes to pMSCs further suggested cultured pMSCs originate from podoplanin+CD36+ cells. This highlights the importance of detailed cell phenotyping to optimise therapeutic capacity, and ensure use of relevant cells in functional assays.

A Cd9+Cd271+ stem/progenitor population and the SHP2 pathway contribute to neonatal-to-adult switching that regulates tendon maturation.

Tendon maturation lays the foundation for postnatal tendon development, its proper mechanical function, and regeneration, but the critical cell populations and the entangled mechanisms remain poorly understood. Here, by integrating the structural, mechanical, and molecular properties, we show that post-natal days 7-14 are the crucial transitional stage for mouse tendon maturation. We decode the cellular and molecular regulatory networks at the single-cell level. We find that a nerve growth factor (NGF)-secreting Cd9+Cd271+ tendon stem/progenitor cell population mainly prompts conversion from neonate to adult tendon. Through single-cell gene regulatory network analysis, in vitro inhibitor identification, and in vivo tendon-specific Shp2 deletion, we find that SHP2 signaling is a regulator for tendon maturation. Our research comprehensively reveals the dynamic cell population transition during tendon maturation, implementing insights into the critical roles of the maturation-related stem cell population and SHP2 signaling pathway during tendon differentiation and regeneration.

CD271 antibody-functionalized microspheres capable of selective recruitment of reparative endogenous stem cells for in situ bone regeneration.

In the strategy of in situ bone regeneration, it used to be difficult to specifically recruit bone marrow mesenchymal stem cells (BM-MSCs) by a single marker. Recently, CD271 has been considered to be one of the most specific markers to isolate BM-MSCs; however, the effectiveness of CD271 antibodies in recruiting BM-MSCs has not been explored yet. In this study, we developed novel CD271 antibody-functionalized chitosan (CS) microspheres with the aid of polydopamine (PDA) coating to recruit endogenous BM-MSCs for in situ bone regeneration. The CS microspheres were sequentially modified with PDA and CD271 antibody through dopamine self-polymerization and bioconjugation, respectively. In vitro studies showed that the CD271 antibody-functionalized microspheres selectively captured significantly more BM-MSCs from a fluorescently labeled heterotypic cell population than non-functionalized controls. In addition, the PDA coating was critical for supporting stable adhesion and proliferation of the captured BM-MSCs. Effective early recruitment of CD271+ stem cells by the functionalized microspheres at bone defect site of SD rat was observed by the CD271/DAPI immunofluorescence staining, which led to significantly enhanced new bone formation in rat femoral condyle defect over long term. Together, findings from this study have demonstrated, for the first time, that the CD271 antibody-functionalized CS microspheres are promising for in situ bone regeneration.

CD271 promotes STZ-induced diabetic wound healing and regulates epidermal stem cell survival in the presence of the pTrkA receptor.

Diabetes mellitus (DM) often causes delayed wound healing in patients, which can lead to limb loss, disability, and even death. Many conventional therapeutic strategies have been proposed, but there is still no effective therapy for DM wounds. This study aimed to explore the effects of CD271 and phosphorylated tyrosine kinase receptor A (pTrkA) on the migration and proliferation abilities of epidermal stem cells (eSCs) and on the activation of DM wound healing. We investigated the interventional effects of CD271-overexpressing eSC (CD271 eSC) treatment and pTrkA inhibition (through k252a treatment) on delayed wound healing using mice with streptozotocin-induced DM. The migration and proliferation abilities of control eSCs, CD271 eSCs, and k252a-treated CD271 eSCs were observed under high-glucose conditions. Decreases in CD271 and increases in pTrkA were observed in DM mouse skin compared with control mouse skin; in addition, the rate of wound closure in DM mice was promoted by CD271 eSC treatment but delayed by pTrkA inhibition. Furthermore, the CD271 eSC migration and proliferation were greater than of control eSCs. Compared with that of CD271 eSCs, the number of CD271+k252a eSCs decreased significantly under high-glucose conditions. In parallel, the expression levels of the pERK, pAkt, and pJNK pathways increased in CD271 eSCs and decreased in CD271+k252a eSCs under high glucose. Our findings demonstrate that CD271 and pTrkA affect DM wound closure by promoting the eSC migration and proliferation. This mechanism involving the pERK, pAkt, and pJNK pathways might be a new therapeutic target for the treatment of delayed wound re-epithelialization in DM.

Characterization of neural crest-derived stem cells isolated from human bone marrow for improvement of transplanted islet function.

Background: Murine boundary cap-derived neural crest stem cells (NCSCs) are capable of enhancing islet function by stimulating beta cell proliferation as well as increasing the neural and vascular density in the islets both in vitro and in vivo. This study aimed to isolate NCSC-like cells from human bone marrow.Methods: CD271 magnetic cell separation and culture techniques were used to purify a NCSC-enriched population of human bone marrow. Analyses of the CD271+ and CD271- fractions in terms of protein expression were performed, and the capacity of the CD271+ bone marrow cells to form 3-dimensional spheres when grown under non-adherent conditions was also investigated. Moreover, the NCSC characteristics of the CD271+ cells were evaluated by their ability to migrate toward human islets as well as human islet-like cell clusters (ICC) derived from pluripotent stem cells.Results: The CD271+ bone marrow population fulfilled the criterion of being multipotent stem cells, having the potential to differentiate into glial cells, neurons as well as myofibroblasts in vitro. They had the capacity to form 3-dimensional spheres as well as an ability to migrate toward human islets, further supporting their NCSC identity. Additionally, we demonstrated similar migration features toward stem cell-derived ICC.Conclusion: The results support the NCSC identity of the CD271-enriched human bone marrow population. It remains to investigate whether the human bone marrow-derived NCSCs have the ability to improve transplantation efficacy of not only human islets but stem cell-derived ICC as well.

Adapalene inhibits ovarian cancer ES-2 cells growth by targeting glutamic-oxaloacetic transaminase 1.

Glutamic-oxaloacetic transaminase 1 (GOT1) regulates cellular metabolism through coordinating the utilization of carbohydrates and amino acids to meet nutrient requirements for sustained proliferation. As such, the GOT1 inhibitor may provide a new strategy for the treatment of various cancers. Adapalene has been approved by FDA for the treatment of acne, pimples and pustules, and it may also contribute to the adjunctive therapy for advanced stages of liver and colorectal cancers. In this work, we first examined the enzyme inhibition of over 500 compounds against GOT1 in vitro. As a result, Adapalene effectively inhibited GOT1 enzyme in a non-competitive manner. MST and DARTS assay further confirmed the high affinity between Adapalene and GOT1. Furthermore, the growth and migration of ovarian cancer ES-2 cells were obviously inhibited by the treatment of Adapalene. And it induced the apoptosis of ES-2 cells according to Western blot and Hoechst 33258 straining. In addition, molecular docking demonstrated that Adapalene coordinated in an allosteric site of GOT1 with low binding energy. Furthermore, knockdown of GOT1 in ES-2 cells decreased their anti-proliferative sensitivity to Adapalene. Together, our data strongly suggest Adapalene, as a GOT1 inhibitor, could be regarded as a potential drug candidate for ovarian cancer therapy.

Intracellular Notch1 Signaling in Cancer-Associated Fibroblasts Dictates the Plasticity and Stemness of Melanoma Stem/Initiating Cells.

Cancer stem cells (CSCs) play critical roles in cancer initiation, metastasis, recurrence, and drug resistance. Recent studies have revealed involvement of cancer-associated fibroblasts (CAFs) in regulating CSCs. However, the intracellular molecular mechanisms that determine the regulatory role of CAFs in modulating the plasticity of CSCs remain unknown. Here, we uncovered that intracellular Notch1 signaling in CAFs serves as a molecular switch, which modulates tumor heterogeneity and aggressiveness by inversely controlling stromal regulation of the plasticity and stemness of CSCs. Using mesenchymal stem cell-derived fibroblasts (MSC-DF) harboring reciprocal loss-of-function and gain-of-function Notch1 signaling, we found that MSC-DFNotch1-/- prompted cocultured melanoma cells to form more spheroids and acquire the phenotype (CD271+ and Nestin+ ) of melanoma stem/initiating cells (MICs), whereas MSC-DFN1IC+/+ suppressed melanoma cell sphere formation and mitigated properties of MICs. MSC-DFNotch1-/- increased stemness of CD271+ MIC, which resultantly exhibited stronger aggressiveness in vitro and in vivo, by upregulating Sox2/Oct4/Nanog expression. Consistently, when cografted with melanoma cells into NOD scid gamma (NSG) mice, MSC-DFNotch1-/- increased, but MSC-DFN1IC+/+ decreased, the amounts of CD271+ MIC in melanoma tissue. The amounts of CD271+ MIC regulated by MSC-DF carrying high or low Notch1 pathway activity is well correlated with capability of melanoma metastasis, supporting that melanoma metastasis is MIC-mediated. Our data demonstrate that intracellular Notch1 signaling in CAFs is a molecular switch dictating the plasticity and stemness of MICs, thereby regulating melanoma aggressiveness, and therefore that targeting the intracellular Notch1 signaling pathway in CAFs may present a new therapeutic strategy for melanoma. Stem Cells 2019;37:865-875.

Angiogenic Potential of Bone Marrow Derived CD133+ and CD271+ Intramyocardial Stem Cell Trans- Plantation Post MI.

BACKGROUND: Bone marrow (BM)-derived stem cells with their various functions and characteristics have become a well-recognized source for the cell-based therapies. However, knowledge on their therapeutic potential and the shortage for a cross-link between distinct BM-derived stem cells, primed after the onset of myocardial infarction (MI), seems to be still rudimentary. Therefore, the post-examination of the therapeutic characteristics of such primed hematopoietic CD133+ and mesenchymal CD271+ stem cells was the object of the present study. METHODS AND RESULTS: The effects of respective CD133+ and CD271+ mononuclear cells alone as well as in the co-culture model have been explored with focus on their angiogenic potential. The phenotypic analysis revealed a small percentage of isolated cells expressing both surface markers. Moreover, target stem cells isolated with our standardized immunomagnetic isolation procedure did not show any negative alterations following BM storage in regard to cell numbers and/or quality. In vitro network formation relied predominantly on CD271+ stem cells when compared with single CD133+ culture. Interestingly, CD133+ cells contributed in the tube formation, only if they were cultivated in combination with CD271+ cells. Additional to the in vitro examination, therapeutic effects of the primed stem cells were investigated 48 h post MI in a murine model. Hence, we have found a lower expression of transforming growth factor ßeta 3 (TGFß3) as well as an increase of the proangiogenic factors after CD133+ cell treatment in contrast to CD271+ cell treatment. On the other hand, the CD271+ cell therapy led to a lower expression of the inflammatory cytokines. CONCLUSION: The interactions between CD271+ and CD133+ subpopulations the extent to which the combination may enhance cardiac regeneration has still not been investigated so far. We expect that the multiple characteristics and various regenerative effects of CD271+ cells alone as well as in combination with CD133+ will result in an improved therapeutic impact on ischemic heart disease.

Low-affinity Nerve Growth Factor Receptor (CD271) Heterogeneous Expression in Adult and Fetal Mesenchymal Stromal Cells.

Human multipotent mesenchymal stromal cells (MSC) are isolated from a plethora of tissue sources for cell therapy purposes. In 2006, the International Society for Cellular Therapy (ISCT) published minimal guidelines to define MSC identity. Nevertheless, many independent studies demonstrated that cells meeting the ISCT criteria possessed heterogeneous phenotypes and functionalities, heavily influenced by culture conditions. In this study, human MSC derived from many adult (bone marrow and adipose tissue) or fetal (cord blood, Wharton's jelly, umbilical cord perivascular compartment and amniotic fluid) tissues were investigated. Their immunophenotype was analyzed to define consistent source-specific markers by extensive flow cytometry analysis and real-time qRT-PCR. CD271+ subpopulations were detected in adult MSC, whereas NG2 was significantly more expressed in fetal MSC but failed validation on independent samples coming from an external laboratory. The highest number of CD271+ adult MSC were detected soon after isolation in serum-based culture conditions. Furthermore, heterogeneous percentages of CD271 expression were found in platelet lysate-based or serum-free culture conditions. Finally, CD271+ adult MSC showed high clonogenic and osteogenic properties as compared to CD271- cells. To conclude, in this phenotype-function correlation study CD271+ subpopulation confers heterogeneity on adult MSC, confirming the need of more specific markers to address MSC properties.

Delivery of adapalene using a novel topical gel based on tea tree oil nano-emulsion: Permeation, antibacterial and safety assessments.

The aim of present study was to design and optimize 0.1% adapalene loaded nano-emulsion to improve the drug efficacy and increase its user compliance. Effect of type and concentration of surfactants was studied on size of 0.1% adapalene loaded nano-emulsion. Optimized formulation was then evaluated for particle size, polydispersity index, morphology, viscosity, and pH. Subsequently, 1% carbopol® 934 was incorporated to the optimized formulation for preparation of its gel form. The efficacy and safety of 0.1% adapalene loaded nano-emulsion gel was assessed compared to marketed gel containing 0.1% adapalene. In-vitro studies showed that adapalene permeation through the skin was negligible in both adapalene loaded nano-emulsion gel and adapalene marketed gel. Furthermore, drug distribution studies in skin indicated higher retention of adapalene in the dermis in adapalene loaded nano-emulsion gel compared with adapalene marketed gel. Antibacterial activity against Propionibacterium acnes showed that adapalene loaded nano-emulsion is effective in reducing minimum inhibitory concentration of the formulation in comparison with tea tree oil nano-emulsion, and pure tea tree oil. In vivo skin irritation studies showed absence of irritancy for adapalene loaded nano-emulsion gel. Also, blood and liver absorption of the drug, histological analysis of liver and liver enzyme activity of rats after 90 days' treatment were investigated. No drug was detected in blood/liver which in addition to an absence of any adverse effect on liver and enzymes showed the potential of adapalene loaded nano-emulsion gel as a novel carrier for topical delivery of adapalene.

The convergent roles of CD271/p75 in neural crest-derived melanoma plasticity.

The embryonic microenvironment is an important source of signals that promote multipotent cells to adopt a specific fate and direct cells along distinct migratory pathways. Yet, the ability of the embryonic microenvironment to retain multipotent progenitors or reprogram de-differentiated cells is less clear. Mistakes in cell differentiation or migration often result in developmental defects and tumorigenesis, including aggressive cancers that share many characteristics with embryonic progenitor cells. This is a striking feature of the vertebrate neural crest, a multipotent and highly migratory cell population first identified by His (1868) with the potential to metamorphose into aggressive melanoma cancer. In this perspective, we address the roles of CD271/p75 in tumor initiation, phenotype switching and reprogramming of metastatic melanoma and discuss the convergence of these roles in melanoma plasticity.

Multiple Autologous Bone Marrow-Derived CD271+ Mesenchymal Stem Cell Transplantation Overcomes Drug-Resistant Epilepsy in Children.

There is a need among patients suffering from drug-resistant epilepsy (DRE) for more efficient and less toxic treatments. The objective of the present study was to assess the safety, feasibility, and potential efficacy of autologous bone marrow cell transplantation in pediatric patients with DRE. Two females and two males (11 months to 6 years) were enrolled and underwent a combined therapy consisting of autologous bone marrow nucleated cells (BMNCs) transplantation (intrathecal: 0.5 × 109 ; intravenous: 0.38 × 109 -1.72 × 109 ) followed by four rounds of intrathecal bone marrow mesenchymal stem cells (BMMSCs) transplantation (18.5 × 106 -40 × 106 ) every 3 months. The BMMSCs used were a unique population derived from CD271-positive cells. The neurological evaluation included magnetic resonance imaging, electroencephalography (EEG), and cognitive development assessment. The characteristics of BMMSCs were evaluated. Four intravenous and 20 intrathecal transplantations into the cerebrospinal fluid were performed. There were no adverse events, and the therapy was safe and feasible over 2 years of follow-up. The therapy resulted in neurological and cognitive improvement in all patients, including a reduction in the number of epileptic seizures (from 10 per day to 1 per week) and an absence of status epilepticus episodes (from 4 per week to 0 per week). The number of discharges on the EEG evaluation was decreased, and cognitive improvement was noted with respect to reactions to light and sound, emotions, and motor function. An analysis of the BMMSCs' characteristics revealed the expression of neurotrophic, proangiogenic, and tissue remodeling factors, and the immunomodulatory potential. Our results demonstrate the safety and feasibility of BMNCs and BMMSCs transplantations and the considerable neurological and cognitive improvement in children with DRE. Stem Cells Translational Medicine 2018;7:20-33.

The feasibility of the CD271+ and CD271- mesenchymal stromal cell enrichment toward nucleus pulposus-like cells.

INTRODUCTION: Factors promoting nerve cell ingrowth are considered responsible for chronic back pain resulting from the intervertebral disc degeneration (IDD). One of the recent exploratory IDD treatments is stem cell transplantation therapy. The CD271 (low-affinity nerve growth factor receptor) has been identified as a mark-er of the most homogeneous mesenchymal stem cell (MSC) subset. It is capable of promoting differentiation along adipogenic, osteogenic and chondrogenic lineages and producing significantly higher levels of cytokines as compared to the total population of plastic adherence-mesenchymal stem cells (PA-MSCs). We investigated the ability of CD271+ MSCs to differentiate into chondrocyte-like cells of the nucleus pulposus (NP) of intervertebral disc. We also examined CD271- MSCs, using PA-MSCs as a control cell population. MATERIAL AND METHODS: Bone marrow derived PA-MSCs and its two subsets, CD271- MSCs and CD271+ MSCs, were seeded in collagen scaffolds. After two weeks of growth in NP-differentiation medium, RNA was isolated from cells-scaffold constructs and was analyzed by q-PCR for expression of NP markers. Glycosaminoglycans were analyzed biochemically directly in cells-scaffold constructs. RESULTS: Expression of NP markers - extracellular matrix components such as aggrecan, collagen type II and glycosaminoglycans on both RNA and the protein levels - was significantly higher in CD271- MSCs compared to the CD271+ MSCs and PA-MSCs cell populations. CONCLUSIONS: CD271- MSCs may be superior candidates for NP restorative treatment compared to CD271+ MSCs and PA-MSCs due to their ability of expressing NP-supporting extracellular matrix components at levels higher than the other two studied MSC subsets.

Intramyocardial fate and effect of iron nanoparticles co-injected with MACS® purified stem cell products.

BACKGROUND: Magnetic activated cell sorting (MACS®) is routinely used to isolate stem cell subpopulations intended for the treatment of cardiovascular diseases. In strong contrast, studies examining the amount, effect and intramyocardial distribution of iron nanoparticles used for magnetic cell labelling are missing, although iron excess can cause functional disorders in the heart. METHODS AND RESULTS: CD133+ haematopoietic and CD271+ mesenchymal stem cells were purified from bone marrow using automatically and manually MACS® based systems. Flow cytometric measurements demonstrated a rapid loss of MACS® MicroBeads from cells under culture conditions, while storage under hypothermic conditions decelerated their detachment. Moreover, an average loading of ∼11 fg iron/cell caused by magnetic labelling was determined in magnetic particle spectroscopy. Importantly, hemodynamic measurements as well as histological examinations using a myocardial ischemia/reperfusion mouse model showed no influence of MACS® MicroBeads on cardiac regeneration, while the transplantation of stem cells caused a significant improvement. Furthermore, immunostainings demonstrated the clearance of co-injected iron nanoparticles from stem cells and the surrounding heart tissue within 48 h post transplantation. CONCLUSIONS: Our results indicate that iron amounts typically co-injected with MACS® purified stem cells do not harm cardiac functions and are cleared from heart tissue within a few hours. Therefore, we conclude that MACS® MicroBeads exhibit a good compatibility in the cardiac environment.

Dermal CD271+ Cells are Closely Associated with Regeneration of the Dermis in the Wound Healing Process.

Stem cells have recently been shown to play important roles in wound healing. The aim of this study was to investigate the role of dermal CD271+ cells in wound healing. Full-thickness wounds were produced on the backs of 5-year-old and 24-week-old mice, and time-course of wound closure, CD271+ cell counts, and gene expression levels were compared. Delayed wound healing was observed in 24-week-old mice. The peak of CD271+ cell increase was delayed in 24-week-old mice, and gene expression levels of growth factors in wounded tissue were significantly increased in 5-year-old mice. Dermal CD271+ cells purified by fluorescence-activated cell sorting (FACS) expressed higher growth factors than CD271- cells, suggesting that CD271+ cells play important roles by producing growth factors. This study also investigated dermal CD271+ cells in patients with chronic skin ulcers. Dermal CD271+ cells in patients were significantly reduced compared with in healthy controls. Thus, dermal CD271+ cells are closely associated with wound healing.