From Mesenchymal Stromal Cells to Engineered Extracellular Vesicles: A New Therapeutic Paradigm.

Mesenchymal stromal cells (MSCs) are multipotent cells obtained from many tissues including bone marrow, adipose tissue, umbilical cord, amniotic fluid, and placenta. MSCs are the leading cell source for stem cell therapy due to their regenerative and immunomodulatory properties, their low risk of tumorigenesis and lack of ethical constraints. However, clinical applications of MSCs remain limited. MSC therapeutic development continues to pose challenges in terms of preparation, purity, consistency, efficiency, reproducibility, processing time and scalability. Additionally, there are issues with their poor engraftment and survival in sites of disease or damage that limit their capacity to directly replace damaged cells. A key recent development in MSC research, however, is the now widely accepted view that MSCs primarily exert therapeutic effects via paracrine factor secretion. One of the major paracrine effectors are extracellular vesicles (EVs). EVs represent a potential cell-free alternative to stem cell therapy but are also rapidly emerging as a novel therapeutic platform in their own right, particularly in the form of engineered EVs (EEVs) tailored to target a broad range of clinical indications. However, the development of EVs and EEVs for therapeutic application still faces a number of hurdles, including the establishment of a consistent, scalable cell source, and the development of robust GMP-compliant upstream and downstream manufacturing processes. In this review we will highlight the clinical challenges of MSC therapeutic development and discuss how EVs and EEVs can overcome the challenges faced in the clinical application of MSCs.

Late/post-term decidual basalis-derived mesenchymal stem/stromal cells show evidence of advanced ageing and downregulation of microRNA-516b-5p.

INTRODUCTION: The placenta is a short-lived organ, yet it shows signs of progressive ageing in the third trimester. Studies of ageing chorionic placental tissue have recently flourished, providing evidence of advanced ageing of tissues in the late/post-term (L/PT) period of gestation. However, ageing of the maternal aspect of the maternal-fetal interface, specifically the decidua basalis, is poorly understood. Here, we investigated whether the L/PT period was associated with advanced ageing and exhaustion of important decidua basalis mesenchymal stem/stromal cells (DMSCs) functions. METHODS: In this study, DMSCs were isolated and characterised from early term (ET) and L/PT placental tissue and they were then investigated by employing various MSC potency and ageing assays. RNA sequencing was also performed to screen for specific microRNAs that are associated with stem cell exhaustion and ageing between ET- and L/PT-DMSCs. RESULTS: L/PT-DMSCs, when compared to ET-DMSCs, showed significantly lower cell proliferation and a significant higher level of cell apoptosis. L/PT-DMSCs showed significantly lower resistance to oxidative stress and a significant decrease in antioxidant capacity compared with ET-DMSCs. Western blot analysis revealed increased expression of the stress-mediated P-p38MAPK protein in L/PT-DMSCs. RNA Sequencing showed microRNA (miR) miR-516b-5p, was present at significantly lower levels in L/PT-DMSCs. Inhibition of miR-516b-5p in ET-DMSCs revealed a decline in the ability of the inhibited cells to survive in extended cell culture. DISCUSSION: These data provide the first evidence of advanced ageing and exhaustion of important stem cell functions in L/PT-DMSCs, and the involvement of specific miRs in the DMSC ageing process.

Functional changes in decidual mesenchymal stem/stromal cells are associated with spontaneous onset of labour.

Ageing and parturition share common pathways, but their relationship remains poorly understood. Decidual cells undergo ageing as parturition approaches term, and these age-related changes may trigger labour. Mesenchymal stem/stromal cells (MSCs) are the predominant stem cell type in the decidua. Stem cell exhaustion is a hallmark of ageing, and thus ageing of decidual MSCs (DMSCs) may contribute to the functional changes in decidual tissue required for term spontaneous labour. Here, we determine whether DMSCs from patients undergoing spontaneous onset of labour (SOL-DMSCs) show evidence of ageing-related functional changes compared with those from patients not in labour (NIL-DMSCs), undergoing Caesarean section. Placentae were collected from term (37-40 weeks of gestation), SOL (n = 18) and NIL (n = 17) healthy patients. DMSCs were isolated from the decidua basalis that remained attached to the placenta after delivery. DMSCs displayed stem cell-like properties and were of maternal origin. Important cell properties and lipid profiles were assessed and compared between SOL- and NIL-DMSCs. SOL-DMSCs showed reduced proliferation and increased lipid peroxidation, migration, necrosis, mitochondrial apoptosis, IL-6 production and p38 MAPK levels compared with NIL-DMSCs (P < 0.05). SOL- and NIL-DMSCs also showed significant differences in lipid profiles in various phospholipids (phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol, phosphatidylserine), sphingolipids (ceramide, sphingomyelin), triglycerides and acyl carnitine (P < 0.05). Overall, SOL-DMSCs had altered lipid profiles compared with NIL-DMSCs. In conclusion, SOL-DMSCs showed evidence of ageing-related reduced functionality, accumulation of cellular damage and changes in lipid profiles compared with NIL-DMSCs. These changes may be associated with term spontaneous labour.

Ageing in human parturition: impetus of the gestation clock in the decidua†.

Despite sharing many common features, the relationship between ageing and parturition remains poorly understood. The decidua is a specialized lining of endometrial tissue, which develops in preparation for pregnancy. The structure and location of the decidua support its role as the physical scaffold for the growing embryo and placenta, and thus, it is vital to sustain pregnancy. Approaching term, the physical support properties of the decidua are naturally weakened to permit parturition. In this review, we hypothesize that the natural weakening of decidual tissue at parturition is promoted by the ageing process. Studies of the ageing-related functional and molecular changes in the decidua at parturition are reviewed and classified using hallmarks of ageing as the framework. The potential roles of decidual mesenchymal stem/stromal cell (DMSC) ageing in labor are also discussed because, although stem cell exhaustion is also a hallmark of ageing, its role in labor is not completely understood. In addition, the potential roles of extracellular vesicles secreted by DMSCs in labor, and their parturition-related miRNAs, are reviewed to gain further insight into this research area. In summary, the literature supports the notion that the decidua ages as the pregnancy progresses, and this may facilitate parturition, suggesting that ageing is the probable impetus of the gestational clocks in the decidua. This conceptual framework was developed to provide a better understanding of the natural ageing process of the decidua during parturition as well as to encourage future studies of the importance of healthy ageing for optimal pregnancy outcomes.

Low-dose aspirin treatment enhances the adhesion of preeclamptic decidual mesenchymal stem/stromal cells and reduces their production of pro-inflammatory cytokines.

Preeclampsia (PE) is a hypertensive disorder of human pregnancy. Low-dose aspirin (acetylsalicylic acid) (60-150 mg/day) is used to prevent PE when taken early in pregnancy. The effect of aspirin on term PE remains uncertain. Abnormal placentation is a hallmark of PE and leads to increased placental oxidative stress, which triggers the release of anti-angiogenic factors that cause local damage to the decidual vasculature. The damage subsequently spreads systemically and culminates in maternal clinical symptoms. Decidua basalis mesenchymal stem/stromal cells (DMSCs) reside in a vascular microenvironment. In PE, DMSCs are exposed to abnormally high levels of oxidative stress and circulating inflammatory factors from the maternal blood. We previously showed that colony-forming unit ability and resistance to oxidative stress in DMSCs are reduced in MSCs derived from term PE pregnancies (PE-DMSCs). The action, if any, of aspirin on term PE-DMSCs has not been reported. In this study, aspirin (5 µg/mL) was found to significantly increase PE-DMSC adhesion compared to untreated PE-DMSCs and gestation-matched control DMSCs (p value < 0.001) but had no effect on PE-DMSC proliferation. ELISA analysis showed that aspirin significantly decreased the production of inflammatory cytokines IFN-γ (p value < 0.05) and IL-8 (p value < 0.001) in PE-DMSCs. In addition, aspirin treatment increased the antioxidant capacity of PE-DMSCs compared with the untreated group (p value < 0.05). This study is the first to reveal a novel, beneficial action of aspirin on PE-DMSCs from term PE pregnancies by improving their adhesion, suppressing their production of pro-inflammatory cytokines production, and increasing their antioxidant capacity. KEY MESSAGES: Preeclampsia (PE) is a serious hypertensive disorder of pregnancy. The risk of PE is reduced by aspirin but the mechanism is poorly understood. Decidua basalis mesenchymal stem/stromal cells (DMSCs) are abnormal in PE. Aspirin treatment improves multiple functions of PE-DMSCs. Improved DMSC function may contribute to the beneficial effect of aspirin.

Circular RNAs: Isolation, characterization and their potential role in diseases.

Circular RNA (circRNA) generated by alternative splicing represents a special class of non-coding RNA molecule. CircRNAs are abundant in the eukaryotic cell cytoplasm and have a characteristic organization, timing of action and disease specificity. In contrast to linear RNA, circRNAs are resistant to RNA exonuclease. Consequently, circRNA escapes normal RNA turnover and this improves circRNA stability. CircRNAs can be degraded by microRNA (miRNA) and this results in linearization of the circRNA, which can then act as competitor to endogenous RNA. Through interactions with disease-related miRNA, circRNA can play an important regulatory role in specific diseases. Furthermore, circRNAs have significant potential to become new clinical diagnostic markers.

The application of decellularized human term fetal membranes in tissue engineering and regenerative medicine (TERM).

Tissue engineering and regenerative medicine (TERM) is a field that applies biology and engineering principles to "restore, maintain or repair a tissue after injury". Besides the potential to treat various diseases, these endeavours increase our understanding of fundamental cell biology. Although TERM has progressed rapidly, engineering a whole organ is still beyond our skills, primarily due to the complexity of tissues. Material science and current manufacturing methods are not capable of mimicking this complexity. Therefore, many researchers explore the use of naturally derived materials that maintain important biochemical, structural and mechanical properties of tissues. Consequently, employing non-cellular components of tissues, particularly the extracellular matrix, has emerged as an alternative to synthetic materials. Because of their complexity, decellularized tissues are not as well defined as synthetic materials but they provide cells with a microenvironment that resembles their natural niche. Decellularized tissues are produced from a variety of sources, among which the fetal membranes are excellent candidates since their supply is virtually unlimited, they are readily accessible with minimum ethical concerns and are often discarded as a biological waste. In this review, we will discuss various applications of decellularized fetal membranes as substrates for the expansion of stem cells, their use as two and three-dimensional scaffolds for tissue regeneration, and their use as cell delivery systems. We conclude that fetal membranes have great potential for use in TERM.

Neuroinflammation pathways: a general review.

Activated microglial cells play an important role in immune and inflammatory responses in central nervous system and neurodegenerative diseases. Many pro-apoptotic pathways are mediated by signaling molecules that are produced during neuroinflammation. In glial cells, NF-κB, a transcription factor, initiates and regulates the expression of several inflammatory processes during inflammation which are attributed to the pathology of the several neurodegenerative diseases. In this review, we discuss the most important neuroinflammatory mediators with their pathways. Attenuating cytokines production and controlling microglial inflammatory response, which are the result of understanding neuroinflammation pathways, are considered therapeutic strategies for treating neurodegenerative diseases with an inflammatory component.

An Update on Inflamm-Aging: Mechanisms, Prevention, and Treatment.

Inflamm-aging is a challenging and promising new branch of aging-related research fields that includes areas such as immunosenescence. Increasing evidence indicates that inflamm-aging is intensively associated with many aging diseases, such as Alzheimer's disease, atherosclerosis, heart disease, type II diabetes, and cancer. Mounting studies have focused on the role of inflamm-aging in disease progression and many advances have been made in the last decade. However, the underlying mechanisms by which inflamm-aging affects pathological changes and disease development are still unclear. Here, we review studies of inflamm-aging that explore the concept, pathological features, mechanisms, intervention, and the therapeutic strategies of inflamm-aging in disease progression.

The Emerging Role of HMGB1 in Neuropathic Pain: A Potential Therapeutic Target for Neuroinflammation.

Neuropathic pain (NPP) is intolerable, persistent, and specific type of long-term pain. It is considered to be a direct consequence of pathological changes affecting the somatosensory system and can be debilitating for affected patients. Despite recent progress and growing interest in understanding the pathogenesis of the disease, NPP still presents a major diagnostic and therapeutic challenge. High mobility group box 1 (HMGB1) mediates inflammatory and immune reactions in nervous system and emerging evidence reveals that HMGB1 plays an essential role in neuroinflammation through receptors such as Toll-like receptors (TLR), receptor for advanced glycation end products (RAGE), C-X-X motif chemokines receptor 4 (CXCR4), and N-methyl-D-aspartate (NMDA) receptor. In this review, we present evidence from studies that address the role of HMGB1 in NPP. First, we review studies aimed at determining the role of HMGB1 in NPP and discuss the possible mechanisms underlying HMGB1-mediated NPP progression where receptors for HMGB1 are involved. Then we review studies that address HMGB1 as a potential therapeutic target for NPP.

Harnessing the secretome of cardiac stem cells as therapy for ischemic heart disease.

Adult stem cells continue to promise opportunities to repair damaged cardiac tissue. However, precisely how adult stem cells accomplish cardiac repair, especially after ischemic damage, remains controversial. It has been postulated that the clinical benefit of adult stem cells for cardiovascular disease results from the release of cytokines and growth factors by the transplanted cells. Studies in animal models of myocardial infarction have reported that such paracrine factors released from transplanted adult stem cells contribute to improved cardiac function by several processes. These include promoting neovascularization of damaged tissue, reducing inflammation, reducing fibrosis and scar formation, as well as protecting cardiomyocytes from apoptosis. In addition, these factors might also stimulate endogenous repair by activating cardiac stem cells. Interestingly, stem cells discovered to be resident in the heart appear to be functionally superior to extra-cardiac adult stem cells when transplanted for cardiac repair and regeneration. In this review, we discuss the therapeutic potential of cardiac stem cells and how the proteins secreted from these cells might be harnessed to promote repair and regeneration of damaged cardiac tissue. We also highlight how recent controversies about the efficacy of adult stem cells in clinical trials of ischemic heart disease have not dampened enthusiasm for the application of cardiac stem cells and their paracrine factors for cardiac repair: the latter have proved superior to the mesenchymal stem cells used in most clinical trials in the past, some of which appear to have been conducted with sub-optimal rigor.

Promoting effect of small molecules in cardiomyogenic and neurogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

Small molecules, growth factors, and cytokines have been used to induce differentiation of stem cells into different lineages. Similarly, demethylating agents can trigger differentiation in adult stem cells. Here, we investigated the in vitro differentiation of rat bone marrow mesenchymal stem cells (MSCs) into cardiomyocytes by a demethylating agent, zebularine, as well as neuronal-like cells by ß-mercaptoethanol in a growth factor or cytokines-free media. Isolated bone marrow-derived MSCs cultured in Dulbecco's Modified Eagle's Medium exhibited a fibroblast-like morphology. These cells expressed positive markers for CD29, CD44, and CD117 and were negative for CD34 and CD45. After treatment with 1 µM zebularine for 24 hours, the MSCs formed myotube-like structures after 10 days in culture. Expression of cardiac-specific genes showed that treated MSCs expressed significantly higher levels of cardiac troponin-T, Nkx2.5, and GATA-4 compared with untreated cells. Immunocytochemical analysis showed that differentiated cells also expressed cardiac proteins, GATA-4, Nkx 2.5, and cardiac troponin-T. For neuronal differentiation, MSCs were treated with 1 and 10 mM ß-mercaptoethanol overnight for 3 hours in complete and serum-free Dulbecco's Modified Eagle's Medium, respectively. Following overnight treatment, neuron-like cells with axonal and dendritic-like projections originating from the cell body toward the neighboring cells were observed in the culture. The mRNA expression of neuronal-specific markers, Map2, Nefl, Tau, and Nestin, was significantly higher, indicating that the treated cells differentiated into neuronal-like cells. Immunostaining showed that differentiated cells were positive for the neuronal markers Flk, Nef, Nestin, and ß-tubulin.

Cardiac Repair With a Novel Population of Mesenchymal Stem Cells Resident in the Human Heart.

Cardiac resident stem cells (CRSCs) hold much promise to treat heart disease but this remains a controversial field. Here, we describe a novel population of CRSCs, which are positive for W8B2 antigen and were obtained from adult human atrial appendages. W8B2(+) CRSCs exhibit a spindle-shaped morphology, are clonogenic and capable of self-renewal. W8B2(+) CRSCs show high expression of mesenchymal but not hematopoietic nor endothelial markers. W8B2(+) CRSCs expressed GATA4, HAND2, and TBX5, but not C-KIT, SCA-1, NKX2.5, PDGFRα, ISL1, or WT1. W8B2(+) CRSCs can differentiate into cardiovascular lineages and secrete a range of cytokines implicated in angiogenesis, chemotaxis, inflammation, extracellular matrix remodeling, cell growth, and survival. In vitro, conditioned medium collected from W8B2(+) CRSCs displayed prosurvival, proangiogenic, and promigratory effects on endothelial cells, superior to that of other adult stem cells tested, and additionally promoted survival and proliferation of neonatal rat cardiomyocytes. Intramyocardial transplantation of human W8B2(+) CRSCs into immunocompromised rats 1 week after myocardial infarction markedly improved cardiac function (∼40% improvement in ejection fraction) and reduced fibrotic scar tissue 4 weeks after infarction. Hearts treated with W8B2(+) CRSCs showed less adverse remodeling of the left ventricle, a greater number of proliferating cardiomyocytes (Ki67(+) cTnT(+) cells) in the remote region, higher myocardial vascular density, and greater infiltration of CD163(+) cells (a marker for M2 macrophages) into the border zone and scar regions. In summary, W8B2(+) CRSCs are distinct from currently known CRSCs found in human hearts, and as such may be an ideal cell source to repair myocardial damage after infarction.

Anticancer and antitumor potential of fucoidan and fucoxanthin, two main metabolites isolated from brown algae.

Seaweed is one of the largest producers of biomass in marine environment and is a rich arsenal of active metabolites and functional ingredients with valuable beneficial health effects. Being a staple part of Asian cuisine, investigations on the crude extracts of Phaeophyceae or brown algae revealed marked antitumor activity, eliciting a variety of research to determine the active ingredients involved in this potential. The sulfated polysaccharide of fucoidan and carotenoid of fucoxanthin were found to be the most important active metabolites of brown algae as potential chemotherapeutic or chemopreventive agents. This review strives to provide detailed account of all current knowledge on the anticancer and antitumor activity of fucoidan and fucoxanthin as the two major metabolites isolated from brown algae.