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The recently emerged novel coronavirus, "severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)", caused a highly contagious disease called coronavirus disease 2019 (COVID-19). It has severely damaged the world's most developed countries and has turned into a major threat for low- and middle-income countries. Since its emergence in late 2019, medical interventions have been substantial, and most countries relied on public health measures collectively known as nonpharmaceutical interventions.To centralize the accumulative knowledge on non-pharmaceutical interventions (NPIs) against COVID-19 for each country under one worldwide consortium.International COVID-19 Research Network collaborators developed a cross-sectional online-survey to assess the implications of NPIs and sanitary supply on incidence and mortality of COVID-19. Survey was conducted between January 1 and February 1, 2021, and participants from 92 countries/territories completed it. The association between NPIs, sanitation supplies and incidence and mortality were examined by multivariate regression, with log-transformed value of population as an offset value.Majority of countries/territories applied several preventive strategies including social distancing (100.0%), quarantine (100.0%), isolation (98.9%), and school closure (97.8%). Individual-level preventive measures such as personal hygiene (100.0%) and wearing facial mask (94.6% at hospital; 93.5% at mass transportation; 91.3% in mass gathering facilities) were also frequently applied. Quarantine at a designated place was negatively associated with incidence and mortality compared to home quarantine. Isolation at a designated place was also associated with reduced mortality compared to home isolation. Recommendations to use sanitizer for personal hygiene reduced incidence compared to recommendation to use soap did. Deprivation of mask was associated with increased incidence. Higher incidence and mortality were found in countries/territories with higher economic level. Mask deprivation was pervasive regardless of economic level.NPIs against COVID-19 such as using sanitizer, quarantine, and isolation can decrease incidence and mortality of COVID-19. This article is protected by copyright. All rights reserved.

Mesenchymal stem cells (MSCs) as a therapeutic promise are often quickly cleared by innate immune cells of the host including natural killer (NK) cells. Efforts have been made to generate immune-escaping human embryonic stem cells (hESCs) where T cell immunity is evaded by defecting β-2-microglobulin (B2M), a common unit for human leukocyte antigen (HLA) class I, and NK cells are inhibited via ectopic expression of HLA-E or -G. However, NK subtypes vary among recipients and even at different pathologic statuses. It is necessary to dissect and optimize the efficacy of the immune-escaping cells against NK subtypes. Here, we first generated B2M knockout hESCs and differentiated them to MSCs (EMSCs) and found that NK resistance occurred with B2M-/- EMSCs expressing HLA-E and -G only when they were transduced via an inducible lentiviral system in a dose-dependent manner but not when they were inserted into a safe harbor. HLA-E and -G expressed at high levels together in transduced EMSCs inhibited three major NK subtypes, including NKG2A+/LILRB1+, NKG2A+/LILRB1-, and NKG2A-/LILRB1+, which was further potentiated by IFN-γ priming. Thus, this study engineers MSCs with resistance to multiple NK subtypes and underscores that dosage matters when a transgene is used to confer a novel effect to host cells, especially for therapeutic cells to evade immune rejection.

The aim of this study is to provide a more accurate representation of COVID-19's case fatality rate (CFR) by performing meta-analyses by continents and income, and by comparing the result with pooled estimates. We used multiple worldwide data sources on COVID-19 for every country reporting COVID-19 cases. On the basis of data, we performed random and fixed meta-analyses for CFR of COVID-19 by continents and income according to each individual calendar date. CFR was estimated based on the different geographical regions and levels of income using three models: pooled estimates, fixed- and random-model. In Asia, all three types of CFR initially remained approximately between 2.0% and 3.0%. In the case of pooled estimates and the fixed model results, CFR increased to 4.0%, by then gradually decreasing, while in the case of random-model, CFR remained under 2.0%. Similarly, in Europe, initially, the two types of CFR peaked at 9.0% and 10.0%, respectively. The random-model results showed an increase near 5.0%. In high-income countries, pooled estimates and fixed-model showed gradually increasing trends with a final pooled estimates and random-model reached about 8.0% and 4.0%, respectively. In middle-income, the pooled estimates and fixed-model have gradually increased reaching up to 4.5%. in low-income countries, CFRs remained similar between 1.5% and 3.0%. Our study emphasizes that COVID-19 CFR is not a fixed or static value. Rather, it is a dynamic estimate that changes with time, population, socioeconomic factors, and the mitigatory efforts of individual countries.

The Coronavirus disease-19 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus -2 (SARS-CoV-2), has impacted human lives in the most profound ways with millions of infections and deaths. Scientists and pharmaceutical companies have been in race to produce vaccines against SARS-CoV-2. Vaccine generation usually demands years of developing and testing for efficacy and safety. However, it only took less than one year to generate two mRNA vaccines from their development to deployment. The rapid production time, cost-effectiveness, versatility in vaccine design, and clinically proven ability to induce cellular and humoral immune response have crowned mRNA vaccines with spotlights as most promising vaccine candidates in the fight against the pandemic. In this review, we discuss the general principles of mRNA vaccine design and working mechanisms of the vaccines, and provide an up-to-date summary of pre-clinical and clinical trials on seven anti-COVID-19 mRNA candidate vaccines, with the focus on the two mRNA vaccines already licensed for vaccination. In addition, we highlight the key strategies in designing mRNA vaccines to maximize the expression of immunogens and avoid intrinsic innate immune response. We also provide some perspective for future vaccine development against COVID-19 and other pathogens.

COVID-19 is hopefully approaching its end in many countries as herd immunity develops and weaker strains of SARS-CoV-2 dominate. However, a new concern occurs over the long-term effects of COVID-19, collectively called "Long COVID", as some symptoms of the nervous system last even after patients recover from COVID-19. This review focuses on studies of anosmia

We have previously demonstrated that mesenchymal stromal/stem cells (MSCs) in spheroids (MSCsubsp/sub) tolerate ambient and hypoxic conditions for a prolonged time. Local administration of MSCsubsp/sub, but not dissociated MSCs (MSCsubdiss/sub), promotes wound healing and relieves multiple sclerosis and osteoarthritis in mice and monkeys. These findings indicate an advantage of MSCsubsp/subover MSCsubdiss/subin sustaining cell viability and efficacy following transplantation, which, however, does not appear to apply to intravenous (i.v.) injection for the principal concern that MSCsubsp/submight cause embolism in small blood vessels of the host, leading to sudden death. Here, we addressed this concern by injecting human MSCsubsp/sub(∼450 μm) or MSCsubdiss/subi.v. into cynomolgus monkeys. Surprisingly, no deaths occurred until sacrifice at day 21 or 60 post injection, and no remarkable physiological changes were found in the animals following the i.v. injection. The big diameters of large blood vessels in monkeys, compared to small animals like mice, may allow sufficient time for MSCsubsp/subto dissociate into single cells so they can pass through small vessels without causing embolism. Retention of MSCsubsp/subwas lower in the lungs but higher in the blood than retention of MSCsubdiss/subat 1 h post injection and both disappeared at day 21. In vitro, MSCsubsp/subtolerated fluidic shear stress with higher survival than MSCsubdiss/sub. Thus, i.v. injection of MSCsubsp/subinto nonhuman primates is feasible, safe, and probably associated with better survival, less lung entrapment and higher efficacy than administration of MSCsubdiss/sub.

Keratinocyte is the predominant cell type in the epidermis of skin, and it provides the protective barrier function for the body. Various signaling pathways have been implicated in keratinocyte differentiation in animal models; However, their temporal regulation and interactions are still to be explored in pluripotent stem cell models. In this report, we use human embryonic stem cells to demonstrate that epidermal ectoderm and subsequent keratinocyte cell fate can be determined step by step under the regulation of defined factors. The inhibition of TGFβ initiates ectodermal lineage differentiation, and the activation of BMP pathway drives epidermal TP63 expression. Meanwhile, the timely activation of WNT pathway suppresses extraembryonic lineage, and promotes epidermal cell fate. With further specification by NOTCH inhibition, more than 90% of cells become TP63-positive stage Ⅱ keratinocytes. Finally, stage Ⅲ keratinocytes are produced under defined hypo-calcium keratinocyte culture conditions, and are further matured in mouse xenograft model. This study not only establishes an in vitro platform to study keratinocyte cell fate determination, but also provides an efficient protocol to produce keratinocytes for disease models and clinical applications.

The outbreak of Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2), has thus far killed over 3,000 people and infected over 80,000 in China and elsewhere in the world, resulting in catastrophe for humans. Similar to its homologous virus, SARS-CoV, which caused SARS in thousands of people in 2003, SARS-CoV-2 might also be transmitted from the bats and causes similar symptoms through a similar mechanism. However, COVID-19 has lower severity and mortality than SARS but is much more transmissive and affects more elderly individuals than youth and more men than women. In response to the rapidly increasing number of publications on the emerging disease, this article attempts to provide a timely and comprehensive review of the swiftly developing research subject. We will cover the basics about the epidemiology, etiology, virology, diagnosis, treatment, prognosis, and prevention of the disease. Although many questions still require answers, we hope that this review helps in the understanding and eradication of the threatening disease.

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.

The genesis of the hair follicle relies on signals derived from mesenchymal cells in the dermis during skin morphogenesis and regeneration. Multipotent skin-derived precursors (SKPs), which exhibit long term proliferation potential when being cultured in spheroids, have been shown to induce hair genesis and hair follicle regeneration in mice, implying a therapeutic potential of SKPs in hair follicle regeneration and bioengineering. However, the hair-inductive property of SKPs declines progressively upon ex vivo culture expansion, suggesting that the expressions of the genes responsible for hair induction are epigenetically unstable. In this study, we found that TSA markedly alleviated culture expansion induced SKP senescence, increased the expression and activity of alkaline phosphatase (AP) in the cells and importantly restored the hair inductive capacity of SKPs. TSA increased the acetylation level of histone H3, including the K19/14 sites in the promoter regions of bone morphogenetic proteins (BMPs) genes, which were associated with elevated gene expression and BMP signaling activity, suggesting a potential attribution of BMP pathway in TSA induced recovery of the hair inductive capacity of SKPs.

The differentiation and maturation of mesenchymal stem cells (MSCs) to mesodermal and other lineages are known to be controlled by various extrinsic and intrinsic signals. The dysregulation of the MSC differentiation balance has been linked to several pathophysiological conditions, including obesity and osteoporosis. Previous research of the molecular mechanisms governing MSC differentiation has mostly focused on transcriptional regulation. However, recent findings are revealing the underrated role of alternative splicing (AS) in MSC differentiation and functions. In this review, we discuss recent progress in elucidating the regulatory roles of AS in MSC differentiation. We catalogue and highlight the key AS events that modulate MSC differentiation to major osteocytes, chondrocytes, and adipocytes, and discuss the regulatory mechanisms by which AS is regulated., Mesenchymal stem cells (MSCs) are essential cell sources for bone, fat tissues, cartilage, and other connective tissues inside the body and in the test tube. Alternative splicing of pre‐mRNAs, which encode critical determinants of MSC differentiation, regulates the MSC differentiation by specifying cell type‐specific transcriptome or antagonizing the function of the WT protein.

Nonhuman primate experimental autoimmune encephalomyelitis (EAE) is a valuable model for multiple sclerosis, an inflammatory demyelinating disease in the central nervous system (CNS). Human embryonic stem cell-derived mesenchymal stem cells (EMSC) are effective in treating murine EAE. Yet, it remains unknown whether the EMSC efficacy is translatable to humans. Here we induced a primate EAE model in cynomolgus monkeys and delivered EMSC in spheres (EMSCsp) to preserve the cell viability during long-distance transportation. EMSCsp intrathecally injected into the CNS, remarkably reduced the clinical symptoms, brain lesions, and neuronal demyelination in the EAE monkeys during a 3-month observation. Whereas, symptoms in the vehicle control-injected EAE monkey remained and reduced slowly and MRI lesions in brain expanded. Moreover, EMSC could transdifferentiate into neural cells in vivo in the CNS of the treated animals. Supporting evidence demonstrated that EMSCsp cells cultured in cerebrospinal fluid from the EAE monkeys largely converted to neural cells with elevated expression of genes for neuronal markers, neurotrophic factors, and neuronal myelination. Thus, this study demonstrates that EMSCsp injected directly into the CNS, can attenuate the disease progression in the primate EAE model, highly encouraging for clinical translation.

Paired box protein 6 (PAX6) is a master regulator of the eye development. Over the last past two decades, our understanding of eye development, especially the molecular function of PAX6, has focused on transcriptional control of the Pax6 expression. However, other regulatory mechanisms for gene expression, including alternative splicing (AS), have been understudied in the eye development. Recent findings suggest that two PAX6 isoforms generated by AS of Pax6 pre-mRNA may play previously underappreciated role(s) during eye development, especially, the corneal development.

Mesenchymal stem cells (MSC) have been derived from a variety of tissues, and cultured either in animal serum-containing (SC) or serum-free (SF) media. We have previously derived MSC from human embryonic stem cells via an intermediate trophoblast step (named EMSC), which also have immunosuppressive and therapeutic effects on animal models of autoimmune disease. To promote the clinical application of this new source of MSC, we report here EMSC derived and cultured in a SF medium MesenCult (SF-EMSC) in comparison with a SC medium (SC-EMSC). SF-EMSC derived in MesenCult also expressed typical MSC markers CD73, CD90, and CD105, and manifested multipotency to differentiate to osteocytes, chondrocytes, and adipocytes. Comparably, CD105+ cells reached 90% about one week slower in the SF than SC conditions, and the proliferation rate was slightly faster for SF-EMSC than SC-EMSC at later passages. Both SF- and SC-EMSC responded similarly to the inflammatory stimulus IFNγ. However, the inflammatory cytokines IL-6 and IL-8 were expressed much less in SF-EMSC than SC-EMSC. Furthermore, knockdown of P16INK4A in both SF- and SC-EMSC reduced replicative senescence. Together, our results suggest that EMSC can be generated in a complete SF condition, and SF-EMSC are largely similar to SC-EMSC. However, it takes longer time to derive EMSC in the SF than SC conditions, and the SF-EMSC proliferate faster at later passages and produce less of the inflammatory cytokines IL-6 and IL-8 than SC-EMSC. This study provides important information for production of clinically applicable EMSC.

Human stem cells are vulnerable to unfavorable conditions, and their transportation relies on costly and inconvenient cryopreservation. We report here that human mesenchymal stem cells (MSC) in spheroids survived ambient conditions (AC) many days longer than in monolayer. Under AC, the viability of MSC in spheroids remained >90% even after seven days, whereas MSC in monolayer mostly died fast. AC-exposed MSC spheroids, after recovery under normal monolayer culture conditions with controlled carbon dioxide and humidity contents, resumed typical morphology and proliferation, and retained differentiating and immunosuppressive capabilities. RNA-sequencing and other assays demonstrate that reduced cell metabolism and proliferation correlates to the enhanced survival of AC-exposed MSC in spheroids versus monolayer. Moreover, AC-exposed MSC, when injected as either single cells or spheroids, retained therapeutic effects in vivo in mouse colitis models. Spheroidal formation also prolonged survival and sustained pluripotency of human embryonic stem cells kept under AC. Therefore, this work offers an alternative and relatively simple method termed spheropreservation versus the conventional method cryopreservation. It shall remarkably simplify long-distance transportation of stem cells of these and probably also other types within temperature-mild areas, and facilitate therapeutic application of MSC as spheroids without further processing.

It has been demonstrated that mesenchymal stem cells (MSCs) differentiated from human embryonic stem cells (hESCs), name EMSCs, can treat a variety of autoimmune and inflammatory diseases, with similar efficacies to those achieved with MSCs derived from somatic tissues such as bone marrow (BMSCs). The chance increases even higher for EMSCs, than somatic tissue derived MSCs​, to become a cell drug as the former can be produced in large scale from an unlimited hESC line with easier quality control and less biosafety concern. We have further demonstrated that both human ESCs and EMSCs, after aggregation to form spheroids, can tolerate hypoxic and ambient conditions (AC) for over 4 and 10 days, respectively, without loss of their viability and alteration of their functions. Based on these advantages, we decided to test whether EMSC spheroids, made in large quantity and delivered through a long-term distance at AC, can treat osteoarthritis spontaneously developed in rhesus macaques (M. mulatta) monkeys as well as the allogenic MSCs. Methods: Xenogeneic AC-transported EMSC spheroids or allogenic BMSCs were injected into the articular cavity of both knees of the monkeys at 3 animals per group. Another two macaques were injected the same way with saline as controls. Results: Both EMSCs and BMSCs groups showed significant amelioration indicated by the reduction of swelling joint size and amplification of keen flare angle post-treatment, compared to the control group. Examinations via X-ray and MRI also indicated the decrease of inflammation and osteophyma, and recovery of the synovium and cartilage in both treated groups. No sign of allergy or graft versus host disease was observed in the animals. Conclusion: Our results demonstrate that human EMSC spheroids can prevent the osteoarthtitis progression and ameliorate osteoarthritis in the rhesus macaques as well as allogenic BMSCs, and this study shall help advance the clinical application of EMSCs.

Mesenchymal stem cells (MSCs) derived from somatic tissues have been used to promote lipotransfer, a common practice in cosmetic surgery. However, the effect of lipotransfer varies, and the mechanism of action remains vague. To address these questions, we differentiated human embryonic stem cells , a stable and unlimited source, into MSCs (EMSCs). Then we subcutaneously transplanted human fat aspirates together with EMSCs or PBS as a control into the back of nude mice . Within 24 h of transplantation, EMSCs promoted aggregation and encapsulation of injected fat tissues. Afterward, all grafts gradually shrank. However, EMSC-containing grafts were larger, heavier and had fewer dark areas on the surface than the control grafts. Histologically, more live adipocytes, vascular cells, and macrophages and less fibrosis were observed in EMSC-containing grafts than in the controls. Some EMSCs differentiated into vascular cells and adipocytes in the EMSC-containing grafts. RNA sequencing revealed that human RNA was shown to decline rapidly, while mouse RNA increased in the grafts; further, human genes related to extracellular matrix remodeling, adipogenesis , and chemokine (including CCL2 ) signaling were expressed at higher levels in the EMSC-containing grafts than they were in the controls. CCL2 knockout reduced macrophage migration towards EMSCs in vitro and early macrophage recruitment to the grafts and the pro-engraftment effect of EMSCs in vivo . Treating mice with a macrophage inhibitor abolished the EMSC effects and converted the grafts to heavy masses of cell debris. Together, these data demonstrate that EMSCs promote fat engraftment via enhanced tissue reconstitution and encapsulation of implanted tissues, which was followed by increased angiogenesis and adipocyte survival and reduced fibrosis, in which stimulated CCL2 signaling and mobilized macrophages play pivotal roles .

Currently, engineered skin substitutes (ESS) are unable to regenerate cutaneous appendages. Recent studies have shown that skin-derived precursors (SKPs), which are extensively available, have the potential to induce hair follicle neogenesis. Here, we demonstrate that ESS consisting of culture-expanded SKPs and epidermal stem cells (Epi-SCs) reconstitute the skin with hair follicle regeneration after grafting into nude mice. SKPs seeded in a C-GAG matrix proliferated and expressed higher levels of hair induction signature genes—such as Akp2, Sox2, CD133 and Bmp6—compared to dermal fibroblasts. Moreover, when ESS prepared by seeding a mixture of culture-expanded murine SKPs and human adult Epi-SCs into a C-GAG matrix was grafted into full-thickness skin wounds in nude mice, black hairs were generated within 3 weeks. Immunofluorescence analysis showed that the SKPs were localized to the dermal papillae of the newly-formed hair follicle. Our results indicate that SKPs can serve as the hair-inductive cells in ESS to furnish it with hair genesis potential

Marfan syndrome (MFS) is a connective tissue disorder caused by mutations in FBN1 gene, which encodes a key extracellular matrix protein FIBRILLIN-1. The haplosufficiency of FBN1 has been implicated in pathogenesis of MFS with manifestations primarily in cardiovascular, muscular, and ocular tissues. Due to limitations in animal models to study the late-onset diseases, human pluripotent stem cells (PSCs) offer a homogeneic tool for dissection of cellular and molecular pathogenic mechanism for MFS in vitro. Here, we first derived induced PSCs (iPSCs) from a MFS patient with a FBN1 mutation and corrected the mutation, thereby generating an isogenic “gain-of-function” control cells for the parental MFS iPSCs. Reversely, we knocked out FBN1 in both alleles in a wild-type (WT) human embryonic stem cell (ESC) line, which served as a loss-of-function model for MFS with the WT cells as an isogenic control. Mesenchymal stem cells derived from both FBN1-mutant iPSCs and -ESCs demonstrated reduced osteogenic differentiation and microfibril formation. We further demonstrated that vascular smooth muscle cells derived from FBN1-mutant iPSCs showed less sensitivity to carbachol as demonstrated by contractility and Ca2+ influx assay, compared to the isogenic controls cells. These findings were further supported by transcriptomic anaylsis of the cells. Therefore, this study based on both gain- and loss-of-function approaches confirmed the pathogenetic role of FBN1 mutations in these MFS-related phenotypic changes.

Despite the long discrepancy over their definition, heterogeneity, and functions, mesenchymal stem cells (MSCs) have proved to be a key player in tissue repair and homeostasis. Generally, somatic tissue-derived MSCs (st-MSCs) are subject to quality variations related to donated samples and biosafety concern for transmission of potential pathogens from the donors. In contrast, human pluripotent stem cells (hPSCs) are unlimited in supply, clear in the biological background, and convenient for quality control, genetic modification, and scale-up production. We, and others, have shown that hPSCs can differentiate in two dimensions or three dimensions to MSCs (ps-MSCs) via embryonic (mesoderm and neural crest) or extraembryonic (trophoblast) cell types under serum-containing or xeno-free and defined conditions. Compared to st-MSCs, ps-MSCs appear less mature, proliferate faster, express lower levels of inflammatory cytokines, and respond less to traditional protocols for st-MSC differentiation to other cell types, especially adipocytes. Nevertheless, ps-MSCs are capable of immune modulation and treatment of an increasing number of animal disease models via mitochondria transfer, paracrine, exosomes, and direct differentiation, and can be potentially used as a universal and endless therapy for clinical application. This review summarizes the progress on ps-MSCs and discusses perspectives and challenges for their potential translation to the clinic. Stem Cells 2019;37:572–581

Liver fibrosis is a disease that causes high morbidity and has become a major health problem. Liver fibrosis can lead to the end stage of liver diseases (livercirrhosisand hepatocellularcarcinoma). Currently, liver transplantation is the only effective treatment for end-stage liver disease. However, the shortage of organ donors, high cost of medical surgery, immunological rejection and transplantation complications severely hamper liver transplantation therapy. Mesenchymal stem cells (MSCs) have been regarded as promising cells for clinical applications in stem cell therapy in the treatment of liver diseases due to their unique multipotent differentiation capacity, immunoregulation and paracrine effects. Although liver fibrosis improvements by MSC transplantation in preclinical experiments as well as clinical trials have been reported, the in vivo fate of MSCs after transportation and their therapeutic mechanisms remain unclear. In this present study, we isolated MSCs from the bone marrow of rhesus macaques. The cells exhibited typical MSC markers and could differentiate into chondrocytes, osteocytes, and adipocytes, which were not affected by labeling with enhanced green fluorescent protein (EGFP). The harvested MSCs respond to interferon-γ stimulation and have the ability to inhibit lymphocyte proliferation in vitro. EGFP-labeled MSCs (1 × 106 cells) were transplanted into mice with carbon tetrachloride-induced liver fibrosis via tail vein injection. The ability of the heterogenic MSC infusion to ameliorate liver fibrosis in mice was evaluated by a blood plasma chemistry index, pathological examination and liver fibrosis-associated gene expression. Additionally, a small number of MSCs that homed and engrafted in the mouse liver tissues were evaluated by immunofluorescence analysis. Our results showed that the transplantation of heterogenic MSCs derived from monkey bone marrow can be used to treat liver fibrosis in the mouse model and that the paracrine effects of MSCs may play an important role in the improvement of liver fibrosis.

Mesenchymal stem/stromal cells (MSCs) have great clinical potential in modulating inflammation and promoting tissue repair. Human embryonic stem cells (hESCs) have recently emerged as a potentially superior cell source for MSCs. However, the generation methods reported so far vary greatly in quality and efficiency. Here, we describe a novel method to rapidly and efficiently produce MSCs from hESCs via a trophoblast-like intermediate stage in approximately 11–16 days. We term these cells “T-MSCs” and show that T-MSCs express a phenotype and differentiation potential minimally required to define MSCs. T-MSCs exhibit potent immunomodulatory activity in vitro as they can remarkably inhibit proliferation of cocultured T and B lymphocytes. Unlike bone marrow MSCs, T-MSCs do not have increased expression of inflammatory mediators in response to IFNγ. Moreover, T-MSCs constitutively express a high level of the immune inhibitory ligand PD-L1 and elicit strong and durable efficacy in two distinct animal models of autoimmune disease, dextran sulfate sodium induced colitis, and experimental autoimmune encephalomyelitis, at doses near those approved for clinical trials. Together, we present a simple and fast derivation method to generate MSCs from hESCs, which possess potent immunomodulatory properties in vitro and in vivo and may serve as a novel and ideal candidate for MSC-based therapies.

Human embryonic stem cell (hESC) derived mesenchymal stem cells (EMSC) are efficacious in treating a series of autoimmune, inflammatory, and degenerative diseases in animal models. However, all the EMSC derivation methods reported so far rely on two-dimensional (2D) culture systems, which are inefficient, costive and difficult for large-scale production. HESC, as an unlimited source, can be successively propagated in spheroids. Here, we demonstrate that hESC spheroids can directly differentiate into MSC spheroids (EMSCSp) within 20 days in one vessel without passaging and the system is scalable to any desired size. EMSCSp can further differentiate into osteocytes and chondrocytes in spheres or demineralized bone matrix. EMSCSp also retains immune-modulatory effects in vitro and therapeutic effects on two mouse models of colitis after dissociation. Compared to EMSC differentiated in monolayer, EMSCSp-derived cells have faster proliferation and higher yield and develop less apoptosis and slower senescence. Thus, the 3D differentiation system allows simple, cost-effective, and scalable production of high-quality EMSC and subsequently bone and cartilage tissues for therapeutic application.

Radiation-induced brain injury (RI) commonly occurs in patients who received head and neck radiotherapy. However, the mechanism of RI remains unclear. We aimed to evaluate whether pyroptosis was involved in RI and the impact of mesenchymal stem cells (MSCs) on it. BALB/c male mice (6-8 weeks) were cranially irradiated (15 Gy), and MSCs were transplanted into the bilateral cortex 2 days later; then mice were sacrificed 1 month later. Meanwhile, irradiated BV-2 microglia cells (10 Gy) were cocultured with MSCs for 24 hours. We observed that irradiated mice brains presented NLRP3 and caspase-1 activation. RT-PCR then indicated that it mainly occurred in microglia cells but not in neurons. Further, irradiated BV-2 cells showed pyroptosis and increased production of IL-18 and IL-1

Radiation-induced brain injury (RI) commonly occurs in patients who received head and neck radiotherapy. However, the mechanism of RI remains unclear. We aimed to evaluate whether pyroptosis was involved in RI and the impact of mesenchymal stem cells (MSCs) on it. BALB/c male mice (6–8 weeks) were cranially irradiated (15 Gy), and MSCs were transplanted into the bilateral cortex 2 days later; then mice were sacrificed 1 month later. Meanwhile, irradiated BV-2 microglia cells (10 Gy) were cocultured with MSCs for 24 hours. We observed that irradiated mice brains presented NLRP3 and caspase-1 activation. RT-PCR then indicated that it mainly occurred in microglia cells but not in neurons. Further, irradiated BV-2 cells showed pyroptosis and increased production of IL-18 and IL-1β. RT-PCR also demonstrated an increased expression of several inflammasome genes in irradiated BV-2 cells, including NLRP3 and AIM2. Particularly, NLRP3 was activated. Knockdown of NLRP3 resulted in decreased LDH release. Noteworthily, in vivo, MSCs transplantation alleviated radiation-induced NLRP3 and caspase-1 activation. Moreover, in vitro, MSCs could decrease caspase-1 dependent pyroptosis, NLRP3 inflammasome activation, and ROS production induced by radiation. Thus, our findings proved that microglia pyroptosis occurred in RI. MSCs may act as a potent therapeutic tool in attenuating pyroptosis.

Ethnopharmacological relevance The Angong Niuhuang Pill (ANP) is a well known Chinese traditional therapeutic for the treatment for diseases affecting the Central Nervous System (CNS). Components of the ANP formulation, including Bovis Calculus Sativus, Pulvis Bubali Comus Concentratus, Moschus , Margarita, Cinnabaris, Realgar, Coptidis Rhizoma, Scutellariae Radix, Gardeniae Fructus, Curcumae Radix, and Bomeolum Syntheticum, have been used for the treatment of stroke, encephalitis and emergency meningitis across Asia, especially in China for hundreds of years. Objective The goal of this study was to investigate the anti-atherosclerosis and cardio-protective effects of ANP administration using a rodent model of atherosclerosis induced by a high fat and vitamin D 3 . Methods Specific Pathogen-Free (SPF) 78 male SD rats were randomly divided into a control group and 5 atherosclerotic model groups. The atherosclerotic groups were divided to receive either Simvastatin (SVTT, 0.005 g/kg), Low-dose ANP (0.125 g/kg), Medium-dose ANP (0.25 g/kg), and High-dose ANP (0.5 g/kg). Following adaptive feeding for one week, atherosclerosis was induced and the atherosclerosis model was established. Experimental drugs (either simvastatin or ANP) or normal saline were administered intragastrically once daily for 9 weeks starting from the 8th week. A carotid artery ultrasound was performed at the 17th week to determine whether atherosclerosis had been induced. After the atherosclerosis model was successfully established, platelet aggregation rates, serum biochemical indices, apoptosis-related Bcl-2, Bax proteins levels in the heart were assayed. Pathological and histological analysis was completed using artery tissue from different experimental different groups to assess the effects of ANP. Results ANP significantly decreased aortic membrane thickness, the maximum platelet aggregation rates, and the ratio of low density lipoprotein cholesterol (LDL) to high density lipoprotein cholesterol (HDL). In addition, ANP significantly reduced serum contents of total cholesterol, low density lipoprotein, malondialdehyde, troponin I, high-sensitivity C-reactive protein, and lactate dehydrogenase. ANP markedly improved abnormal pathological conditions of the aorta and heart, and helped to prevent myocardial apoptosis. Conclusions We have demonstrated that ANP has robust ant-atherosclerosis and cardio-protective effects on a high-fat and vitamin D 3 – induced rodent model of atherosclerosis due to its antiplatelet aggregation, lipid regulatory, antioxidant, anti-inflammatory and anti-apoptotic properties.

Induced pluripotent stem cells (iPSC) hold tremendous potential for personalized cell-based repair strategies to treat musculoskeletal disorders. To establish human iPSCs as a potential source of viable chondroprogenitors for articular cartilage repair, we assessed the in vitro chondrogenic potential of the pluripotent population versus an iPSC-derived mesenchymal-like progenitor population. We found the direct plating of undifferentiated iPSCs into high-density micromass cultures in the presence of BMP-2 promoted chondrogenic differentiation, however these conditions resulted in a mixed population of cells resembling the phenotype of articular cartilage, transient cartilage, and fibrocartilage. The progenitor cells derived from human iPSCs exhibited immunophenotypic features of mesenchymal stem cells (MSCs) and developed along multiple mesenchymal lineages, including osteoblasts, adipocytes, and chondrocytes in vitro. The data indicate the derivation of a mesenchymal stem cell population from human iPSCs is necessary to limit culture heterogeneity as well as chondrocyte maturation in the differentiated progeny. Moreover, as compared to pellet culture differentiation, BMP-2 treatment of iPSC-derived MSC-like (iPSC-MSC) micromass cultures resulted in a phenotype more typical of articular chondrocytes, characterized by the enrichment of cartilage-specific type II collagen (Col2a1), decreased expression of type I collagen (Col1a1) as well as lack of chondrocyte hypertrophy. These studies represent a first step toward identifying the most suitable iPSC progeny for developing cell-based approaches to repair joint cartilage damage.

The efficiency of embryonic stem (ES) cell derivation relies on an optimized culture medium and techniques for treating preimplantation stage embryos. Recently, ES cell derivation from the preblastocyst developmental stage was reported by removing the zona pellucida from embryos of the most efficient strain for ES cell derivation (129Sv) during early preimplantation. Here, we showed that ES cells can be efficiently derived and maintained in a modified medium (MEMα), from preblastocysts of a low-efficiency mouse strain (a hybrid consisting of 50% B6, 25% CBA, and 25% DBA). Preblastocyst-derived ES cell lines were normal in terms of pluripotency-related protein expression, and chromosome number. Also, preblastocyst-derived ES cell lines from various culture conditions showed pluripotency in vivo through teratoma analysis. Interestingly, ES cell lines produced from preblastocysts and blastocysts, regardless of the derivation culture conditions, are nearly indistinguishable by their global gene expression profiles.

The de novo DNA methyltransferase DNMT3B functions in establishing DNA methylation patterns during development. DNMT3B missense mutations cause immunodeficiency, centromere instability and facial anomalies (ICF) syndrome. The restriction of Dnmt3b expression to neural progenitor cells, as well as the mild cognitive defects observed in ICF patients, suggests that DNMT3B may play an important role in early neurogenesis. We performed RNAi knockdown of DNMT3B in human embryonic stem cells (hESCs) in order to investigate the mechanistic contribution of DNMT3B to DNA methylation and early neuronal differentiation. While DNMT3B was not required for early neuroepithelium specification, DNMT3B deficient neuroepithelium exhibited accelerated maturation with earlier expression, relative to normal hESCs, of mature neuronal markers (such as NEUROD1) and of early neuronal regional specifiers (such as those for the neural crest). Genome-wide analyses of DNA methylation by MethylC-seq identified novel regions of hypomethylation in the DNMT3B knockdowns along the X chromosome as well as pericentromeric regions, rather than changes to promoters of specific dysregulated genes. We observed a loss of H3K27me3 and the polycomb complex protein EZH2 at the promoters of early neural and neural crest specifier genes during differentiation of DNMT3B knockdown but not normal hESCs. Our results indicate that DNMT3B mediates large-scale methylation patterns in hESCs and that DNMT3B deficiency in the cells alters the timing of their neuronal differentiation and maturation.