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Currently, more than 30 000 allogeneic hematopoietic stem cell (HSC) transplantations have been performed for the treatment of hematological and nonhematological diseases using HSC from umbilical cord blood (CB). However, the wide utilization of CB as a source of HSC is limited by the low number of cells recovered. One strategy to expand ex vivo CB‐HSC is represented by the use of bone marrow mesenchymal stromal cells (BM‐MSCs) as a feeder to enhance HSC proliferation while maintaining HSC stemness. Indeed, BM‐MSCs have been recognized as one of the most relevant players in the HSC niche. Thus, it has been hypothesized that they can support the ex vivo expansion of HSC by mimicking the physiological microenvironment present in the hematopoietic niche. Due to the role of placenta in supporting fetal hematopoiesis, MSC derived from the amniotic membrane (hAMSC) of human term placenta could represent an interesting alternative to BM‐MSC as a feeder layer to enhance the proliferation and maintain HSC stemness. Therefore, in this study we investigated if hAMSC could support the ex vivo expansion of HSC and progenitor cells. The capacity of hAMSCs to support the ex vivo expansion of CB‐HSC was evaluated in comparison to the control condition represented by the CB‐CD34+ cells without a feeder layer. The coculture was performed at two different CD34+:MSC ratios (1:2 and 1:8) in both cell‐to‐cell contact and transwell setting. After 7 days, the cells were collected and analyzed for phenotype and functionality. Our results suggest that hAMSCs represent a valuable alternative to BM‐MSC to support: (a) the ex vivo expansion of CB‐HSC in both contact and transwell systems, (b) the colony forming unit ability, and (c) long‐term culture initiating cells ability. Overall, these findings may contribute to address the unmet need of high HSC content in CB units available for transplantation., Ex vivo expansion of cord blood hematopoietic stem cell (HSC) and progenitor cell is required to improve efficacy of transplantation in adults. Bone marrow mesenchymal stromal cells are the gold standard for HSC expansion, but they can be difficult to obtain. Amniotic membrane‐derived MSC can be a valid alternative that could help meet the demand of HSC for transplants.

Background It is now well established that factors (free or in extracellular vesicles) secreted by mesenchymal stromal cells (MSC) are important mediators of MSC regenerative actions. Herein we produced the secretome (conditioned medium, CM) from MSC isolated from the amniotic membrane (hAMSC) and CM from the intact amniotic membrane (hAM, no manipulation or enzymatic digestion) in order to potentially identify an effective, easy and less expensive secretome to produce for potential applications in regenerative medicine. Given that immunomodulation is a key mechanism of action through which hAMSC contributes to tissue regeneration, we used a comprehensive panel of in vitro immunomodulatory tests to compare the CMs. Methods Amniotic membranes were either cut into fragments or used for hAMSC isolation. CMs from hAMSC at passages 0 and 2 were collected after a standard 5-day culture while CM from hAM was collected after a 2- and 5-day culture. Immunomodulation was assessed in terms of PBMC and T-cell proliferation, T-cell subset polarization, T-regulatory cell induction, cell cytotoxicity and monocyte differentiation toward antigen-presenting cells. Furthermore, we performed a comparison between CM obtained from single donors and pooled CM. We also assessed the impact of lyophilization on the immunomodulatory properties of CM. Results We demonstrate that CM from hAM has comparable immunomodulatory properties to CM from hAMSC at passages 0 and 2. Furthermore, we demonstrate that pooled CMs have similar effects when compared to CM from single donors used separately. Finally, we demonstrate that lyophilization does not alter the in vitro immunomodulatory properties of CM from hAM and hAMSC. Conclusions The results presented herein support the possibility to produce secretome from intact hAM and open the prospect to highly improve the scalability of the GMP production process while reducing the costs and time related to the process of cell isolation and expansion. Moreover, the possibility of having a lyophilized secretome that maintains its original properties would allow for a ready-to-use product with easier handling, shipping and storage. The use of a lyophilized product will also facilitate clinicians by permitting customized reconstitution volumes and methods according to the most suitable formula required by the clinical application.

Human amniotic membrane and amniotic membrane-derived mesenchymal stromal cells (hAMSCs) have produced promising results in regenerative medicine, especially for the treatment of inflammatory-based diseases and for different injuries including those in the orthopedic field such as tendon disorders. hAMSCs have been proposed to exert their anti-inflammatory and healing potential via secreted factors, both free and conveyed within extracellular vesicles (EVs). In particular, EV miRNAs are considered privileged players due to their impact on target cells and tissues, and their future use as therapeutic molecules is being intensely investigated. In this view, EV-miRNA quantification in either research or future clinical products has emerged as a crucial paradigm, although, to date, largely unsolved due to lack of reliable reference genes (RGs). In this study, a panel of thirteen putative miRNA RGs (let-7a-5p, miR-16-5p, miR-22-5p, miR-23a-3p, miR-26a-5p, miR-29a-5p, miR-101-3p, miR-103a-3p, miR-221-3p, miR-423-5p, miR-425-5p, miR-660-5p and U6 snRNA) that were identified in different EV types was assessed in hAMSC-EVs. A validated experimental pipeline was followed, sifting the output of four largely accepted algorithms for RG prediction (geNorm, NormFinder, BestKeeper and &Delta, Ct method). Out of nine RGs constitutively expressed across all EV isolates, miR-101-3p and miR-22-5p resulted in the most stable RGs, whereas miR-423-5p and U6 snRNA performed poorly. miR-22-5p was also previously reported to be a reliable RG in adipose-derived MSC-EVs, suggesting its suitability across samples isolated from different MSC types. Further, to shed light on the impact of incorrect RG choice, the level of five tendon-related miRNAs (miR-29a-3p, miR-135a-5p, miR-146a-5p, miR-337-3p, let-7d-5p) was compared among hAMSC-EVs isolates. The use of miR-423-5p and U6 snRNA did not allow a correct quantification of miRNA incorporation in EVs, leading to less accurate fingerprinting and, if used for potency prediction, misleading indication of the most appropriate clinical batch. These results emphasize the crucial importance of RG choice for EV-miRNAs in hAMSCs studies and contribute to the identification of reliable RGs such as miR-101-3p and miR-22-5p to be validated in other MSC-EVs related fields.

Growing evidence suggests a mechanistic link between inflammation and the development and progression of fibrotic processes. Mesenchymal stromal cells derived from the human amniotic membrane (hAMSCs), which display marked immunomodulatory properties, have been shown to reduce bleomycin‐induced lung fibrosis in mice, possibly by creating a microenvironment able to limit the evolution of chronic inflammation to fibrosis. However, the ability of hAMSCs to modulate immune cells involved in bleomycin‐induced pulmonary inflammation has yet to be elucidated. Herein, we conducted a longitudinal study of the effects of hAMSCs on alveolar and lung immune cell populations upon bleomycin challenge. Immune cells collected through bronchoalveolar lavage were examined by flow cytometry, and lung tissues were used to study gene expression of markers associated with different immune cell types. We observed that hAMSCs increased lung expression of T regulatory cell marker Foxp3, increased macrophage polarization toward an anti‐inflammatory phenotype (M2), and reduced the antigen‐presentation potential of macrophages and dendritic cells. For the first time, we demonstrate that hAMSCs markedly reduce pulmonary B‐cell recruitment, retention, and maturation, and counteract the formation and expansion of intrapulmonary lymphoid aggregates. Thus, hAMSCs may hamper the self‐maintaining inflammatory condition promoted by B cells that continuously act as antigen presenting cells for proximal T lymphocytes in injured lungs. By modulating B‐cell response, hAMSCs may contribute to blunting of the chronicization of lung inflammatory processes with a consequent reduction of the progression of the fibrotic lesion., This study demonstrates that amniotic mesenchymal stromal cells reduce pulmonary fibrosis in bleomycin‐challenged mice by creating an anti‐inflammatory microenvironment mediated by their ability to control B‐cell recruitment, retention, maturation, and to reduce the formation/expansion of lymphoid aggregates in diseased lungs.

Chondral and osteochondral lesions represent one of the most challenging problems in the orthopedic field, as these types of injuries lead to disability and worsened quality of life for patients and have an economic impact on the healthcare system. The aim of this in vivo study was to develop a new tissue engineering approach through a hybrid scaffold for osteochondral tissue regeneration made of porous polyurethane foam (PU) coated under vacuum with calcium phosphates (PU/VAC). Scaffold characterization showed a highly porous and interconnected structure. Human amniotic mesenchymal stromal cells (hAMSCs) were loaded into scaffolds using pectin (PECT) as a carrier. Osteochondral defects in medial femoral condyles of rabbits were created and randomly allocated in one of the following groups: plain scaffold (PU/VAC), scaffold with hAMSCs injected in the implant site (PU/VAC/hAMSC), scaffold with hAMSCs loaded in pectin (PU/VAC/PECT/hAMSC), and no treated defects (untreated). The therapeutic efficacy was assessed by macroscopic, histological, histomorphometric, microtomographic, and ultrastructural analyses at 3, 6, 12, and 24 weeks. Histological results showed that the scaffold was permissive to tissue growth and penetration, an immature osteocartilaginous tissue was observed at early experimental times, with a more accentuated bone regeneration in comparison with the cartilage layer in the absence of any inflammatory reaction.

During pregnancy, a successful coexistence between the mother and the semi-allogenic fetus occurs which requires a dynamic immune system to guarantee an efficient immune protection against possible infections and tolerance toward fetal antigens. The mechanism of fetal-maternal tolerance is still an open question. There is growing in vitro and in vivo evidence that mesenchymal stromal cells (MSC) which are present in perinatal tissues have a prominent role in generating a functional microenvironment critical to a successful pregnancy. This review highlights the immunomodulatory properties of perinatal MSC and their impact on the major immune cell subsets present in the uterus during pregnancy, such as natural killer cells, antigen-presenting cells (macrophages and dendritic cells), and T cells. Here, we discuss the current understanding and the possible contribution of perinatal MSC in the establishment of fetal-maternal tolerance, providing a new perspective on the physiology of gestation.

Human term placenta is comprised of various tissues from which different cells can be obtained, including hematopoietic stem cells and mesenchymal stem/stromal cells (MSCs). Areas covered: This review will discuss the possibility to incorporate placental tissue cells in cord blood banking. It will discuss general features of human placenta, with a brief review of the immune cells at the fetal-maternal interface and the different cell populations isolated from placenta, with a particular focus on MSCs. It will address the question as to why placenta-derived MSCs should be banked with their hematopoietic counterparts. It will discuss clinical trials which are studying safety and efficacy of placenta tissue-derived MSCs in selected diseases, and preclinical studies which have proven their therapeutic properties in other diseases. It will discuss banking of umbilical cord blood and raise several issues for improvement, and the applications of cord blood cells in non-malignant disorders. Expert commentary: Umbilical cord blood banking saves lives worldwide. The concomitant banking of non-hematopoietic cells from placenta, which could be applied therapeutically in the future, alone or in combination to their hematopoietic counterparts, could exploit current banking processes while laying the foundation for clinical trials exploring placenta-derived cell therapies in regenerative medicine.

Among the many cell types useful in developing therapeutic treatments, human amniotic cells from placenta have been proposed as valid candidates. Both human amniotic epithelial and mesenchymal stromal cells, and the conditioned medium generated from their culture, exert multiple immunosuppressive activities. Indeed, they inhibit T and B cell proliferation, suppress inflammatory properties of monocytes, macrophages, dendritic cells, neutrophils, and natural killer cells, while promoting induction of cells with regulatory functions such as regulatory T cells and anti-inflammatory M2 macrophages. These properties have laid the foundation for their use for the treatment of inflammatory-based diseases, and encouraging results have been obtained in different preclinical disease models where exacerbated inflammation is present. Moreover, an immune-privileged status of amniotic cells has been often highlighted. However, even if long-term engraftment of amniotic cells has been reported into immunocompetent animals, only few cells survive after infusion. Furthermore, amniotic cells have been shown to be able to induce immune responses in vivo and, under specific culture conditions, they can stimulate T cell proliferation in vitro. Although immunosuppressive properties are a widely recognized characteristic of amniotic cells, immunogenic and stimulatory activities appear to be less reported, sporadic events. In order to improve therapeutic outcome, the mechanisms responsible for the suppressive versus stimulatory activity need to be carefully addressed. In this review, both the immunosuppressive and immunostimulatory activity of amniotic cells will be discussed.

Placenta-derived mesenchymal stromal cells (MSC) have often been considered to linger behind their equivalents from other tissues, such as MSC from bone marrow, in many aspects including their therapeutic potential in regenerative medicine. Nowadays however, it is clear that certain aspects make placental MSC attractive as a cellular therapy, such as their lack of ethical concerns and ease of isolation from human term placenta, a material long regarded as biological waste. Moreover, placental MSC virtually lack expression of human leukocyte antigens and co-stimulatory molecules, making them very attractive for transplantation in allogeneic settings. In the context of cancer, cell therapy remains an area of intense investigation whereby MSC have been shown to play opposing roles, and placental MSC are no exception. In this review, we will discuss dichotomy of placental MSC that underscores the challenges in understanding their therapeutic potential in oncology.

The osteogenic potential of mesenchymal stromal cells (MSCs) varies among different tissue sources. Strontium enhances the osteogenic differentiation of bone marrow-derived MSCs (BM-MSCs), but whether it exerts similar effects on placental decidual basalis-derived MSCs (PDB-MSCs) remains unknown. Here, we compared the influence of strontium on the proliferation and osteogenic differentiation of human PDB- and BM-MSCs in vitro. We found that 1 mM and 10 mM strontium, but not 0.1 mM strontium, evidently promoted the proliferation of human PDB- and BM-MSCs. These doses of strontium showed a comparable alkaline phosphatase activity in both cell types, but their osteogenic gene expressions were promoted in a dose-dependent manner. Strontium at doses of 0.1 mM and 1 mM elevated several osteogenic gene expressions of PDB-MSCs, but not those of BM-MSCs at an early stage. Nevertheless, they failed to enhance the mineralization of either cell type. By contrast, 10 mM strontium facilitated the osteogenic gene expression as well as the mineralization of human PDB- and BM-MSCs. Collectively, this study demonstrated that human PDB- and BM-MSCs shared a great similarity in response to strontium, which promoted their proliferation and osteogenic differentiation in a dose-dependent manner.

Inflammation significantly impacts the progression of Huntington's disease (HD) and the mutant HTT protein determines a pro‐inflammatory activation of microglia. Mesenchymal stem/stromal cells (MSC) from the amniotic membrane (hAMSC), and their conditioned medium (CM‐hAMSC), have been shown to possess protective effects in vitro and in vivo in animal models of immune‐based disorders and of traumatic brain injury, which have been shown to be mediated by their immunomodulatory properties. In this study, in the R6/2 mouse model for HD we demonstrate that mice treated with CM‐hAMSC display less severe signs of neurological dysfunction than saline‐treated ones. CM‐hAMSC treatment significantly delayed the development of the hind paw clasping response during tail suspension, reduced deficits in rotarod performance, and decreased locomotor activity in an open field test. The effects of CM‐hAMSC on neurological function were reflected in a significant amelioration in brain pathology, including reduction in striatal atrophy and the formation of striatal neuronal intranuclear inclusions. In addition, while no significant increase was found in the expression of BDNF levels after CM‐hAMSC treatment, a significant decrease of microglia activation and inducible nitric oxide synthase levels were observed. These results support the concept that CM‐hAMSC could act by modulating inflammatory cells, and more specifically microglia.

Macroautophagy/autophagy occurs at basal levels in all eukaryotic cells and plays an important role in maintaining bio-energetic homeostasis through the control of molecule degradation and organelle turnover. It can be induced by environmental conditions such as starvation, and is deregulated in many diseases including autoimmune diseases, neurodegenerative disorders, and cancer. Interestingly, the modulation of autophagy in mesenchymal stem cells (MSCs) represents a possible mechanism which, affecting MSC properties, may have an impact on their regenerative, therapeutic potential. Furthermore, the ability of MSCs to modulate autophagy of cells in injured tissues/organs has been recently proposed to be involved in the regeneration of damaged tissues and organs. In particular, MSCs can affect autophagy in immune cells involved in injury-induced inflammation reducing their survival, proliferation, and function and favoring the resolution of inflammation. In addition, MSCs can affect autophagy in endogenous adult or progenitor cells, promoting their survival, proliferation and differentiation supporting the restoration of functional tissue. This review provides, for the first time, an overview of the studies which highlight a possible link between the therapeutic properties of MSCs and their ability to modulate autophagy, and it summarizes examples of disorders where these therapeutic properties have been correlated with such modulation. A better elucidation of the mechanism(s) through which MSCs can modulate the autophagy of target cells and how autophagy can affect MSCs therapeutic properties, can provide a wider perspective for the clinical application of MSCs in the treatment of many diseases.Abbreviations: 3-MA: 3-methyladenine; AD: Alzheimer disease; ATG: autophagy-related; BECN1: beclin 1; BM: bone marrow; CD: cluster of differentiation; EAE: experimental autoimmune encephalomyelitis; IL: interleukin; INF: interferon; LAP: LC3-associated phagocytosis; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MSCs: mesenchymal stem cells; MTOR: mechanistic target of rapamycin kinase; PD: Parkinson disease; PtdIns3K: class III phosphatidylinositol 3-kinase; ROS: reactive oxygen species; SLE: systemic lupus erythematosus; SQSTM1: sequestosome 1; TBI: traumatic brain injury; TGF: transforming growth factor; TNF: tumor necrosis factor.

Mesenchymal stromal cells from the human amniotic membrane (i.e., human amniotic mesenchymal stromal cells [hAMSCs]) of term placenta are increasingly attracting attention for their applications in regenerative medicine. Osteochondral defects represent a major clinical problem with lifelong chronic pain and compromised quality of life. Great promise for osteochondral regeneration is held in hydrogel-based constructs that have a flexible composition and mimic the physiological structure of cartilage. Cell loading within a hydrogel represents an advantage for regenerative purposes, but the encapsulation steps can modify cell properties. As pectin gels have also been explored as cell vehicles on 3D scaffolds, the aim of this study was to explore the possibility to include hAMSCs in pectin gel. Immobilization of hAMSCs into pectin gels could expand their application in cell-based bioengineering strategies. hAMSCs were analyzed for their viability and recovery from the pectin gel and for their ability to differentiate toward the osteogenic lineage and to maintain their immunological characteristics. When treated with a purposely designed pectin/hydroxyapatite gel biocomposite, hAMSCs retained their ability to differentiate toward the osteogenic lineage, did not induce an immune response, and retained their ability to reduce T cell proliferation. Taken together, these results suggest that hAMSCs could be used in combination to pectin gels for the study of novel osteochondral regeneration strategies.

Different in vitro and in vivo studies have demonstrated that cells isolated from the amniotic membrane possess broad immunomodulatory activities. Indeed, both amniotic epithelial cells (hAECs) and amniotic mesenchymal stromal cells (hAMSCs) suppress B cell and T cell proliferation, generation of Th1, Th17, and cytotoxic T cells, and production of inflammatory cytokines. Moreover, both hAMSCs and hAECs suppress inflammatory activities of monocytes, macrophages, dendritic cells, neutrophils, and natural killer cells, while promoting the induction of cells with regulatory functions such as regulatory T cells and antiinflammatory M2 macrophages. Increasing evidence shows that these immunomodulatory properties are the mechanism through which amniotic cells can exert the therapeutic effect observed in the preclinical setting of diseases characterized by an exacerbated inflammatory response. In this chapter, we will comprehensively review in vitro and in vivo studies of the immunomodulatory properties of hAECs and hAMSCs and discuss differences between the two cell populations.

Objectives To define the features of human amniotic mesenchymal stromal cell secretome and its protective properties in experimental models of acute brain injury. Design Prospective experimental study. Setting Laboratory research. Subjects C57Bl/6 mice. Interventions Mice subjected to sham or traumatic brain injury by controlled cortical impact received human amniotic mesenchymal stromal cells or phosphate-buffered saline infused intracerebroventricularly or intravenously 24 hours after injury. Organotypic cortical brain slices exposed to ischemic injury by oxygen-glucose deprivation were treated with human amniotic mesenchymal stromal cells or with their secretome (conditioned medium) in a transwell system. Measurements and main results Traumatic brain injured mice receiving human amniotic mesenchymal stromal cells intravenously or intracerebroventricularly showed early and lasting functional and anatomical brain protection. cortical slices injured by oxigen-glucose deprivation and treated with human amniotic mesenchymal stromal cells or conditioned medium showed comparable protective effects (neuronal rescue, promotion of M2 microglia polarization, induction of trophic factors) indicating that the exposure of human amniotic mesenchymal stromal cells to the injured tissue is not necessary for the release of bioactive factors. Using sequential size-exclusion and gel-filtration chromatography, we identified a conditioned medium subfraction, which specifically displays these highly protective properties and we found that this fraction was rich in bioactive molecules with molecular weight smaller than 700 Da. Quantitative RNA analysis and mass spectrometry-based peptidomics showed that the active factors are not proteins or RNAs. The metabolomic profiling of six metabolic classes identified a list of molecules whose abundance was selectively elevated in the active conditioned medium fraction. Conclusions Human amniotic mesenchymal stromal cell-secreted factors protect the brain after acute injury. Importantly, a fraction rich in metabolites, and containing neither proteic nor ribonucleic molecules was protective. This study indicates the profiling of protective factors that could be useful in cell-free therapeutic approaches for acute brain injury.

In the cell therapy scenario, efficient tracing of transplanted cells is essential for investigating cell migration and interactions with host tissues. This is fundamental to provide mechanistic insights which altogether allow for the understanding of the translational potential of placental cell therapy in the clinical setting. Mesenchymal stem/stromal cells (MSC) from human placenta are increasingly being investigated for their potential in treating patients with a variety of diseases. In this study, we investigated the feasibility of using poly (methyl methacrylate) nanoparticles (PMMA‐NPs) to trace placental MSC, namely those from the amniotic membrane (hAMSC) and early chorionic villi (hCV‐MSC). We report that PMMP‐NPs are efficiently internalized and retained in both populations, and do not alter cell morphofunctional parameters. We observed that PMMP‐NP incorporation does not alter in vitro immune modulatory capability of placental MSC, a characteristic central to their reparative/therapeutic effects in vitro. We also show that in vitro, PMMP‐NP uptake is not affected by hypoxia. Interestingly, after in vivo brain ischaemia and reperfusion injury achieved by transient middle cerebral artery occlusion (tMCAo) in mice, iv hAMSC treatment resulted in significant improvement in cognitive function compared to PBS‐treated tMCAo mice. Our study provides evidence that tracing placental MSC with PMMP‐NPs does not alter their in vitro and in vivo functions. These observations are grounds for the use of PMMP‐NPs as tools to investigate the therapeutic mechanisms of hAMSC and hCV‐MSC in preclinical models of inflammatory‐driven diseases., Journal of Cellular and Molecular Medicine, 20 (6), ISSN:1582-1838, ISSN:1582-4934

During the past several years, the human placenta and in particular the amniotic fetal membrane have attracted much attention as a possible source of cells to be used in cell therapy approaches due to its putative stem cell potential associated with its early embryonic origin and its immunomodulatory potential associated with its role in fetomaternal tolerance.Within the human amniotic membrane, it is possible to isolate two main cell populations: amniotic epithelial cells (from the epithelial layer of the amniotic membrane) and amniotic mesenchymal cells (from the mesenchymal layer of the amniotic membrane). In this chapter, we will describe a method for the isolation of mesenchymal stromal cells (hAMSCs). We will also describe their optimal culture conditions and the phenotypic characterization of cells after passaging.

Pre‐eclampsia (PE) is one of the most severe syndromes in human pregnancy, and the underlying mechanisms of PE have yet to be determined. Pre‐eclampsia is characterized by the alteration of the immune system's activation status, an increase in inflammatory Th1/Th17/APC cells, and a decrease in Th2/Treg subsets/cytokines. Moreover, inflammatory infiltrates have been detected in the amniotic membranes of pre‐eclamptic placentae, and to this date limited data are available regarding the role of amniotic membrane cells in PE. Interestingly, we and others have previously shown that human amniotic mesenchymal stromal cells (hAMSC) possess anti‐inflammatory properties towards almost all immune cells described to be altered in PE. In this study we investigated whether the immunomodulatory properties of hAMSC were altered in PE. We performed a comprehensive study of cell phenotype and investigated the in vitro immunomodulatory properties of hAMSC isolated from pre‐eclamptic pregnancies (PE‐hAMSC), comparing them to hAMSC from normal pregnancies (N‐hAMSC). We demonstrate that PE‐hAMSC inhibit CD4/CD8 T‐cell proliferation, suppress Th1/Th2/Th17 polarization, induce Treg and block dendritic cells and M1 differentiation switching them to M2 cells. Notably, PE‐hAMSC generated a more prominent induction of Treg and higher suppression of interferon‐γ when compared to N‐hAMSC, and this was associated with higher transforming growth factor‐β1 secretion and PD‐L2/PD‐L1 expression in PE‐hAMSC. In conclusion, for the first time we demonstrate that there is no intrinsic impairment of the immunomodulatory features of PE‐hAMSC. Our results suggest that amniotic mesenchymal stromal cells do not contribute to the disease, but conversely, could participate in offsetting the inflammatory environment which characterizes PE.

Background In order to shed light on the regenerative mechanism of mesenchymal stem cells (MSCs) in vivo, the bio-distribution profile of implanted cells using a stable and long-term tracking method is needed. We herein investigated the bio-distribution of human placental deciduas basalis derived MSCs (termed as PDB-MSCs) in nude mice after intravenous injection by carbon radioisotope labeling thymidine (14C-TdR), which is able to incorporate into new DNA strands during cell replication. Results The proliferation rate and radioactive emission of human PDB-MSCs after labeled with different concentrations of 14C-TdR were measured. PDB-MSCs labeled with 1 μCi possessed high radioactivity, and the biological characteristics (i.e. morphology, colony forming ability, differentiation capabilities, karyotype and cell cycle) showed no significant changes after labeling. Thus, 1 μCi was the optimal concentration in this experimental design. In nude mice, 1 × 10614C-TdR-labeled PDB-MSCs were injected intravenously and the organs were collected at days 1, 2, 3, 5, 30 and 180 after injection, respectively. Radiolabeled PDB-MSCs were found mainly in the lung, liver, spleen, stomach and left femur of the recipient nude mice at the whole observation period. Conclusions This work provided solid evidence that 14C-TdR labeling did not alter the biological characteristics of human placental MSCs, and that this labeling method has potential to decrease the signal from non-infused or dead cells for cell tracking. Therefore, this labeling technique can be utilized to quantify the infused cells after long-term follow-up in pre-clinical studies.

Mesenchymal stromal cells (MSC) isolated from human term placental tissues possess unique characteristics, including their peculiar immunomodulatory properties and their multilineage differentiation potential. The osteogenic differentiation capacity of placental MSC has been widely disputed, and continues to be an issue of debate. This review will briefly discuss the different MSC populations which can be obtained from different regions of human term placenta, along with their unique properties, focusing specifically on their osteogenic differentiation potential. We will present the strategies used to enhance osteogenic differentiation potential in vitro, such as through the selection of subpopulations more prone to differentiate, the modification of the components of osteo-inductive medium, and even mechanical stimulation. Accordingly, the applications of three-dimensional environments in vitro and in vivo, such as non-synthetic, polymer-based, and ceramic scaffolds, will also be discussed, along with results obtained from pre-clinical studies of placental MSC for the regeneration of bone defects and treatment of bone-related diseases.

Placenta-derived mesenchymal stromal cells (MSC) have attracted more attention for their immune modulatory properties and poor immunogenicity, which makes them suitable for allogeneic transplantation. Although MSC isolated from different areas of the placenta share several features, they also present significant biological differences, which might point to distinct clinical applications. Hence, we compared cells from full term placenta distinguishing them on the basis of their origin, either maternal or fetal. We used cells developed by Pluristem LTD: PLacenta expanded mesenchymal-like adherent stromal cells (PLX), maternal-derived cells (PLX-PAD), fetal-derived cells (PLX-R18), and amniotic membrane-derived MSC (hAMSC). We compared immune modulatory properties evaluating effects on T-lymphocyte proliferation, expression of cytotoxicity markers, T-helper and T-regulatory cell polarization, and monocyte differentiation toward antigen presenting cells (APC). Furthermore, we investigated cell immunogenicity. We show that MSCs and MSC-like cells from both fetal and maternal sources present immune modulatory properties versus lymphoid (T cells) and myeloid (APC) cells, whereby fetal-derived cells (PLX-R18 and hAMSC) have a stronger capacity to modulate immune cell proliferation and differentiation. Our results emphasize the importance of understanding the cell origin and characteristics in order to obtain a desired result, such as modulation of the inflammatory response that is critical in fostering regenerative processes.