Distinct in vitro and in vivo neutralization profiles of monoclonal antibodies elicited by the receptor binding domain of the ancestral SARS-CoV-2.

Broadly neutralizing antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants are sought to curb coronavirus disease 2019 (COVID-19) infections. Here we produced and characterized a set of mouse monoclonal antibodies (mAbs) specific for the ancestral SARS-CoV-2 receptor binding domain (RBD). Two of them, 17A7 and 17B10, were highly potent in microneutralization assay with 50% inhibitory concentration (IC50 ) ≤135 ng/mL against infectious SARS-CoV-2 variants, including G614, Alpha, Beta, Gamma, Delta, Epsilon, Zeta, Kappa, Lambda, B.1.1.298, B.1.222, B.1.5, and R.1. Both mAbs (especially 17A7) also exhibited strong in vivo efficacy in protecting K18-hACE2 transgenic mice from the lethal infection with G614, Alpha, Beta, Gamma, and Delta viruses. Structural analysis indicated that 17A7 and 17B10 target the tip of the receptor binding motif in the RBD-up conformation. A third RBD-reactive mAb (3A6) although escaped by Beta and Gamma, was highly effective in cross-neutralizing Delta and Omicron BA.1 variants in vitro and in vivo. In competition experiments, antibodies targeting epitopes similar to these 3 mAbs were rarely enriched in human COVID-19 convalescent sera or postvaccination sera. These results are helpful to inform new antibody/vaccine design and these mAbs can be useful tools for characterizing SARS-CoV-2 variants and elicited antibody responses.

Impacts of residential indoor air quality and environmental risk factors on adult asthma-related health outcomes in Chicago, IL.

BACKGROUND: Residential environments are known to contribute to asthma. OBJECTIVE: To examine the joint impacts of exposures to residential indoor and outdoor air pollutants and housing risk factors on adult asthma-related health outcomes. METHODS: We analyzed >1-year of data from 53 participants from 41 homes in the pre-intervention period of the Breathe Easy Project prior to ventilation and filtration retrofits. Health outcomes included surveys of asthma control, health-related quality of life, stress, and healthcare utilizations. Environmental assessments included quarterly measurements of indoor and outdoor pollutants (e.g., HCHO, CO, CO2, NO2, O3, and PM), home walk-throughs, and surveys of environmental risk factors. Indoor pollutant concentrations were also matched with surveys of time spent at home to estimate indoor pollutant exposures. RESULTS: Cross-sectional analyses using mixed-effects models indicated that lower annual average asthma control test (ACT) scores were associated (p < 0.05) with higher indoor NO2 (concentration/exposure: ß = -2.42/-1.57), indoor temperature (ß = -1.03 to -0.94), and mold/dampness (ß = -3.09 to -2.41). In longitudinal analysis, lower ACT scores were also associated (p < 0.05) with higher indoor NO2 concentrations (ß = -0.29), PM1 (concentration/exposure: ß = -0.12/-0.24), PM2.5 (concentration/exposure: ß = -0.12/-0.26), and PM10 (concentration/exposure: ß = 10.14/-0.28). Emergency department visits were associated with poorer asthma control [incidence rate ratio (IRR) = 0.84; p < 0.001], physical health (IRR = 0.95; p < 0.05), mental health (IRR = 0.95; p < 0.05), higher I/O NO2 ratios (IRR = 1.30; p < 0.05), and higher indoor temperatures (IRR = 1.41; p < 0.05). SIGNIFICANCE: Findings suggest that residential risk factors, including indoor air pollution (especially NO2 and particulate matter), higher indoor temperature, and mold/dampness, may contribute to poorer asthma control. IMPACT: This study highlights the importance of residential indoor air quality and environmental risk factors for asthma control, health-related quality of life, and emergency department visits for asthma. Two timescales of mixed models suggest that exposure to indoor NO2 and particulate matter, higher indoor temperature, and mold/dampness was associated with poorer asthma control. Additionally, emergency department visits were associated with poorer asthma control and health-related quality of life, as well as higher I/O NO2 ratios and indoor temperatures. These findings deepen our understanding of the interrelationships between housing, air quality, and health, and have important implications for programs and policy.

Enhanced virulence and waning vaccine-elicited antibodies account for breakthrough infections caused by SARS-CoV-2 delta and beyond.

Here we interrogate the factors responsible for SARS-CoV-2 breakthrough infections in a K18-hACE2 transgenic mouse model. We show that Delta and the closely related Kappa variant cause viral pneumonia and severe lung lesions in K18-hACE2 mice. Human COVID-19 mRNA post-vaccination sera after the 2nd dose are significantly less efficient in neutralizing Delta/Kappa than early 614G virus in vitro and in vivo. By 5 months post-vaccination, ≥50% of donors lack detectable neutralizing antibodies against Delta and Kappa and all mice receiving 5-month post-vaccination sera die after the lethal challenges. Although a 3rd vaccine dose can boost antibody neutralization against Delta in vitro and in vivo, the mean log neutralization titers against the latest Omicron subvariants are 1/3-1/2 of those against the original 614D virus. Our results suggest that enhanced virulence, greater immune evasion, and waning of vaccine-elicited protection account for SARS-CoV-2 variants caused breakthrough infections.

Indoor air quality impacts of residential mechanical ventilation system retrofits in existing homes in Chicago, IL.

Mechanical ventilation systems are used in residences to introduce ventilation air and dilute indoor-generated pollutants. A variety of ventilation system types can be used in home retrofits, influencing indoor air quality (IAQ) in different ways. Here we describe the Breathe Easy Project, a >2-year longitudinal, pseudo-randomized, crossover study designed to assess IAQ and adult asthma outcomes before and after installing residential mechanical ventilation systems in 40 existing homes in Chicago, IL. Each home received one of three types of ventilation systems: continuous exhaust-only, intermittent powered central-fan-integrated-supply (CFIS), or continuous balanced system with an energy recovery ventilator (ERV). Homes with central heating and/or cooling systems also received MERV 10 filter replacements. Approximately weeklong field measurements were conducted at each home on a quarterly basis throughout the study to monitor environmental conditions, ventilation operation, and indoor and outdoor pollutants, including size-resolved particles (0.3-10 µm), ozone (O3), nitrogen dioxide (NO2), carbon dioxide (CO2), carbon monoxide (CO), and indoor formaldehyde (HCHO). Mean reductions in indoor/outdoor (I/O) ratios across all systems after the intervention were approximately 12% (p = 0.001), 10% (p = 0.008), 42% (p < 0.001), 39% (p = 0.002), and 33% (p = 0.007), for CO2, NO2, and estimated PM1, PM2.5, and PM10, respectively. There was a reduction in I/O ratios for all measured constituents with each type of system, on average, but with varying magnitude and levels of statistical significance. The magnitudes of mean differences in I/O pollutant concentrations ratios were generally largest for most pollutants in the homes that received continuous balanced with ERV and smallest in the homes that received intermittent CFIS systems, with apparent benefits to providing ventilation continuously rather than intermittently. All ventilation system types maintained similar indoor temperatures during pre- and post-intervention periods.

A human iPSC-derived inducible neuronal model of Niemann-Pick disease, type C1.

BACKGROUND: Niemann-Pick disease, type C (NPC) is a childhood-onset, lethal, neurodegenerative disorder caused by autosomal recessive mutations in the genes NPC1 or NPC2 and characterized by impaired cholesterol homeostasis, a lipid essential for cellular function. Cellular cholesterol levels are tightly regulated, and mutations in either NPC1 or NPC2 lead to deficient transport and accumulation of unesterified cholesterol in the late endosome/lysosome compartment, and progressive neurodegeneration in affected individuals. Previous cell-based studies to understand the NPC cellular pathophysiology and screen for therapeutic agents have mainly used patient fibroblasts. However, these do not allow modeling the neurodegenerative aspect of NPC disease, highlighting the need for an in vitro system that permits understanding the cellular mechanisms underlying neuronal loss and identifying appropriate therapies. This study reports the development of a novel human iPSC-derived, inducible neuronal model of Niemann-Pick disease, type C1 (NPC1). RESULTS: We generated a null i3Neuron (inducible × integrated × isogenic) (NPC1-/- i3Neuron) iPSC-derived neuron model of NPC1. The NPC1-/- and the corresponding isogenic NPC1+/+ i3Neuron cell lines were used to efficiently generate homogenous, synchronized neurons that can be used in high-throughput screens. NPC1-/- i3Neurons recapitulate cardinal cellular NPC1 pathological features including perinuclear endolysosomal storage of unesterified cholesterol, accumulation of GM2 and GM3 gangliosides, mitochondrial dysfunction, and impaired axonal lysosomal transport. Cholesterol storage, mitochondrial dysfunction, and axonal trafficking defects can be ameliorated by treatment with 2-hydroxypropyl-ß-cyclodextrin, a drug that has shown efficacy in NPC1 preclinical models and in a phase 1/2a trial. CONCLUSION: Our data demonstrate the utility of this new cell line in high-throughput drug/chemical screens to identify potential therapeutic agents. The NPC1-/- i3Neuron line will also be a valuable tool for the NPC1 research community to explore the pathological mechanisms contributing to neuronal degeneration.

Graphene Quantum Dots from Carbonized Coffee Bean Wastes for Biomedical Applications.

Recent studies concerning graphene quantum dots (GQDs) focus extensively on their application in biomedicine, exploiting their modifiable optical properties and ability to complex with various molecules via π-π or covalent interactions. Among these nascent findings, the potential therapeutic efficacy of GQDs was reported against Parkinson's disease, which has to date remained incurable. Herein, we present an environmentally friendly approach for synthesizing GQDs through a waste-to-treasure method, specifically from coffee waste to nanodrug. Consistent with the previous findings with carbon fiber-derived GQDs, the inhibitory effects of coffee bean-derived GQDs demonstrated similar effectiveness against abnormal α-synuclein fibrillation and the protection of neurons from relevant subcellular damages. The fact that a GQDs-based nanodrug can be prepared from a non-reusable yet edible source illustrates a potential approach to convert such waste materials into novel therapeutic agents with minimal psychological rejection by patients.

Therapeutic Features and Updated Clinical Trials of Mesenchymal Stem Cell (MSC)-Derived Exosomes.

Identification of the immunomodulatory and regenerative properties of mesenchymal stem cells (MSCs) have made them an attractive alternative therapeutic option for diseases with no effective treatment options. Numerous clinical trials have followed; however, issues such as infusional toxicity and cellular rejection have been reported. To address these problems associated with cell-based therapy, MSC exosome therapy was developed and has shown promising clinical outcomes. MSC exosomes are nanosized vesicles secreted from MSCs and represent a non-cellular therapeutic agent. MSC exosomes retain therapeutic features of the cells from which they originated including genetic material, lipids, and proteins. Similar to MSCs, exosomes can induce cell differentiation, immunoregulation, angiogenesis, and tumor suppression. MSC exosomes have therefore been employed in several experimental models and clinical studies. Here, we review the therapeutic potential of MSC-derived exosomes and summarize currently ongoing clinical trials according to disease type. In addition, we propose several functional enhancement strategies for the effective clinical application of MSC exosome therapy.

Induced neural stem cells from human patient-derived fibroblasts attenuate neurodegeneration in Niemann-Pick type C mice.

BACKGROUND: Niemann-Pick disease type C (NPC) is caused by the mutation of NPC genes, which leads to the abnormal accumulation of unesterified cholesterol and glycolipids in lysosomes. This autosomal recessive disease is characterized by liver dysfunction, hepatosplenomegaly, and progressive neurodegeneration. Recently, the application of induced neural stem cells (iNSCs), converted from fibroblasts using specific transcription factors, to repair degenerated lesions has been considered a novel therapy. OBJECTIVES: The therapeutic effects on NPC by human iNSCs generated by our research group have not yet been studied in vivo; in this study, we investigate those effects. METHODS: We used an NPC mouse model to efficiently evaluate the therapeutic effect of iNSCs, because neurodegeneration progress is rapid in NPC. In addition, application of human iNSCs from NPC patient-derived fibroblasts in an NPC model in vivo can give insight into the clinical usefulness of iNSC treatment. The iNSCs, generated from NPC patient-derived fibroblasts using the SOX2 and HMGA2 reprogramming factors, were transplanted by intracerebral injection into NPC mice. RESULTS: Transplantation of iNSCs showed positive results in survival and body weight change in vivo. Additionally, iNSC-treated mice showed improved learning and memory in behavior test results. Furthermore, through magnetic resonance imaging and histopathological assessments, we observed delayed neurodegeneration in NPC mouse brains. CONCLUSIONS: iNSCs converted from patient-derived fibroblasts can become another choice of treatment for neurodegenerative diseases such as NPC.

Graphene Quantum Dots Alleviate Impaired Functions in Niemann-Pick Disease Type C in Vivo.

While the neuropathological characteristics of Niemann-Pick disease type C (NPC) result in a fatal diagnosis, the development of clinically available therapeutic agent remains a challenge. Here we propose graphene quantum dots (GQDs) as a potential candidate for the impaired functions in NPC in vivo. In addition to the previous findings that GQDs exhibit negligible long-term toxicity and are capable of penetrating the blood-brain barrier, GQD treatment reduces the aggregation of cholesterol in the lysosome through expressed physical interactions. GQDs also promote autophagy and restore defective autophagic flux, which, in turn, decreases the atypical accumulation of autophagic vacuoles. More importantly, the injection of GQDs inhibits the loss of Purkinje cells in the cerebellum while also demonstrating reduced activation of microglia. The ability of GQDs to alleviate impaired functions in NPC proves the promise and potential of the use of GQDs toward resolving NPC and other related disorders.

Graphene quantum dots as anti-inflammatory therapy for colitis.

While graphene and its derivatives have been suggested as a potential nanomedicine in several biomimetic models, their specific roles in immunological disorders still remain elusive. Graphene quantum dots (GQDs) may be suitable for treating intestinal bowel diseases (IBDs) because of their low toxicity in vivo and ease of clearance. Here, GQDs are intraperitoneally injected to dextran sulfate sodium (DSS)-induced chronic and acute colitis model, and its efficacy has been confirmed. In particular, GQDs effectively prevent tissue degeneration and ameliorate intestinal inflammation by inhibiting TH1/TH17 polarization. Moreover, GQDs switch the polarization of macrophages from classically activated M1 to M2 and enhance intestinal infiltration of regulatory T cells (Tregs). Therefore, GQDs effectively attenuate excessive inflammation by regulating immune cells, indicating that they can be used as promising alternative therapeutic agents for the treatment of autoimmune disorders, including IBDs.

cAMP/EPAC Signaling Enables ETV2 to Induce Endothelial Cells with High Angiogenesis Potential.

Although the generation of ETV2-induced endothelial cells (iECs) from human fibroblasts serves as a novel therapeutic strategy in regenerative medicine, the process is inefficient, resulting in incomplete iEC angiogenesis. Therefore, we employed chromatin immunoprecipitation (ChIP) sequencing and identified molecular mechanisms underlying ETV2-mediated endothelial transdifferentiation to efficiently produce iECs retaining appropriate functionality in long-term culture. We revealed that the majority of ETV2 targets in human fibroblasts are related to vasculature development and signaling transduction pathways, including Rap1 signaling. From a screening of signaling pathway modulators, we confirmed that forskolin facilitated efficient and rapid iEC reprogramming via activation of the cyclic AMP (cAMP)/exchange proteins directly activated by cAMP (EPAC)/RAP1 axis. The iECs obtained via cAMP signaling activation showed superior angiogenesis in vivo as well as in vitro. Moreover, these cells could form aligned endothelium along the vascular lumen ex vivo when seeded into decellularized liver scaffold. Overall, our study provided evidence that the cAMP/EPAC/RAP1 axis is required for the efficient generation of iECs with angiogenesis potential.

Disease-specific primed human adult stem cells effectively ameliorate experimental atopic dermatitis in mice.

Although human mesenchymal stem cells (hMSCs) hold considerable promise as an alternative therapeutic reagent for allergic disorders including atopic dermatitis (AD), the strategy for enhancing hMSC-based therapy remains challenging. We sought to investigate whether preconditioning with mast cell (MC) granules could enhance the therapeutic efficiency of human umbilical cord blood-derived MSCs (hUCB-MSCs) against AD. Methods: AD was experimentally induced in NC/Nga mice by repeated applications of 4% sodium dodecyl sulfate (SDS) and dermatophagoides farinae (Df) extract, and preconditioned hUCB-MSCs were subcutaneously injected. The therapeutic effect was determined by gross examination and additional ex vivo experiments performed using blood and skin samples to determine the resolution of allergic inflammation. To explore the underlying mechanisms, several co-culture assays with primary isolated immune cells and wound closure assays were conducted. Results: Pretreatment of MC granules enhanced the therapeutic effects of hUCB-MSCs by attenuating the symptoms of AD in an experimental animal model. MC granule-primed cells suppressed the activation of major disease-inducing cells, MCs and B lymphocytes more efficiently than naïve cells both in vitro and in vivo. Histamine-mediated upregulation of the COX-2 signaling pathway was shown to play a crucial role in suppression of the allergic immune response by MC-pretreated hUCB-MSCs. Moreover, MC pretreatment improved the wound healing ability of hUCB-MSCs. Conclusions: Our findings indicate that pre-exposure to MC granules improved the therapeutic effect of hUCB-MSCs on experimental AD by resolving the allergic immune reaction and accelerating the tissue regeneration process more efficiently than naïve cells, suggesting a potential enhancement strategy for stem cell-based therapy.

Human umbilical cord blood plasma alleviates age-related olfactory dysfunction by attenuating peripheral TNF-α expression.

Social requirements are needed for living in an aging society and individual longevity. Among them, improved health and medical cares, appropriate for an aging society are strongly demanded. Human cord blood-derived plasma (hUCP) has recently emerged for its unique anti-aging effects. In this study, we investigated brain rejuvenation, particularly olfactory function, that could be achieved by a systemic administration of young blood and its underlying mechanisms. Older than 24-month-old mice were used as an aged group and administered with intravenous injection of hUCP repetitively, eight times. Anti-aging effect of hUCP on olfactory function was evaluated by buried food finding test. To investigate the mode of action of hUCP, brain, serum and spleen of mice were collected for further ex vivo analyses. Systemic injection of hUCP improved aging-associated olfactory deficits, reducing time for finding food. In the brain, although an infiltration of activated microglia and its expression of cathepsin S remarkably decreased, significant changes of proinflammatory factors were not detected. Conversely, peripheral immune balance distinctly switched from predominance of Type 1 helper T (Th1) cells to alternative regulatory T cells (Tregs). These findings indicate that systemic administration of hUCP attenuates age-related neuroinflammation and subsequent olfactory dysfunction by modulating peripheral immune balance toward Treg cells, suggesting another therapeutic function and mechanism of hUCP administration. [BMB Reports 2019; 52(4): 259-264].

Interferon-γ-mediated secretion of tryptophanyl-tRNA synthetases has a role in protection of human umbilical cord blood-derived mesenchymal stem cells against experimental colitis.

Mesenchymal stem cells (MSCs) are multipotent adult stem cells that present immunosuppressive effects in experimental and clinical trials targeting various rare diseases including inflammatory bowel disease (IBD). In addition, recent studies have reported tryptophanyl-tRNA synthetase (WRS) possess uncanonical roles such as angiostatic and anti-inflammatory effects. However, little is known about the function of WRS in MSC-based therapy. In this study, we investigated if a novel factor, WRS, secreted from MSCs has a role in amelioration of IBD symptoms and determined a specific mechanism underlying MSC therapy. Experimental colitis was induced by administration of 3% DSS solution to 8-week-old mice and human umbilical cord blood-derived MSCs (hUCB-MSCs) were injected intraperitoneally. Secretion of WRS from hUCB-MSCs and direct effect of WRS on isolated CD4＋ T cells was determined via in vitro experiments and hUCB-MSCs showed significant therapeutic rescue against experimental colitis. Importantly, WRS level in serum of colitis induced mice decreased and recovered by administration of MSCs. Through in vitro examination, WRS expression of hUCB-MSCs increased when cells were treated with interferon-γ (IFN-γ). WRS was evaluated and revealed to have a role in inhibiting activated T cells by inducing apoptosis. In summary, IFN-γ- mediated secretion of WRS from MSCs has a role in suppressive effect on excessive inflammation and disease progression of IBD and brings new highlights in the immunomodulatory potency of hUCB-MSCs. [BMB Reports 2019; 52(5): 318-323].

Stem cell-secreted 14,15- epoxyeicosatrienoic acid rescues cholesterol homeostasis and autophagic flux in Niemann-Pick-type C disease.

We previously demonstrated that the direct transplantation of human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs) into the dentate gyrus ameliorated the neurological symptoms of Niemann-Pick type C1 (NPC1)-mutant mice. However, the clinical presentation of NPC1-mutant mice was not fully understood with a molecular mechanism. Here, we found 14,15-epoxyeicosatrienoic acid (14,15-EET), a cytochrome P450 (CYP) metabolite, from hUCB-MSCs and the cerebella of NPC1-mutant mice and investigated the functional consequence of this metabolite. Our screening of the CYP2J family indicated a dysregulation in the CYP system in a cerebellar-specific manner. Moreover, in Purkinje cells, CYP2J6 showed an elevated expression level compared to that of astrocytes, granule cells, and microglia. In this regard, we found that one CYP metabolite, 14,15-EET, acts as a key mediator in ameliorating cholesterol accumulation. In confirming this hypothesis, 14,15-EET treatment reduced the accumulation of cholesterol in human NPC1 patient-derived fibroblasts in vitro by suppressing cholesterol synthesis and ameliorating the impaired autophagic flux. We show that the reduced activity within the CYP system in the cerebellum could cause the neurological symptoms of NPC1 patients, as 14,15-EET treatment significantly rescued cholesterol accumulation and impaired autophagy. We also provide evidence that the intranasal administration of hUCB-MSCs is a highly promising alternative to traumatic surgical transplantation for NPC1 patients.

MIS416 Enhances Therapeutic Functions of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Against Experimental Colitis by Modulating Systemic Immune Milieu.

Human adult stem cells, including umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs), have recently been considered a promising alternative treatment for inflammatory bowel disease (IBD) due to their unique immunomodulatory properties and ability to promote tissue regeneration. However, despite many years of research and pre-clinical studies, results from clinical trials using these cells have been diverse and conflicting. This discrepancy is caused by several factors, such as poor engraftment, low survival rate, and donor-dependent variation of the cells. Enhancement of consistency and efficacy of MSCs remains a challenge for the feasibility of cell-based therapy. In this study, we investigated whether administration of MIS416, a novel microparticle that activates NOD2 and TLR9 signaling, could enhance the therapeutic efficacy of hUCB-MSCs against Crohn's disease, using dextran sulfate sodium (DSS)-induced colitis model. Colitis was experimentally induced in mice by using 3% DSS, and mice were administered a retro-orbital injection of MIS416 and subsequent intraperitoneal injection of hUCB-MSCs. Mice were examined grossly, and blood, spleen, and colon tissues were subsequently collected for further ex vivo analyses. To explore the effects of MIS416 on the therapeutic process, hUCB-MSCs and primary isolated immune cells were cultured with MIS416, and in vitro assays were performed. Compared to the single administration of hUCB-MSCs, co-administration with MIS416 improved the therapeutic efficiency of the stem cells by significantly alleviating the symptoms of IBD. Interestingly, MIS416 did not exert any direct effect on the immunomodulatory capacity of hUCB-MSCs. Instead, systemically injected MIS416 altered the immune milieu in the colon which caused hUCB-MSCs to be more readily recruited toward the lesion site and to suppress inflammation more efficiently. In addition, considerable numbers of regulatory immune cells were stimulated as a result of the cooperation of MIS416 and hUCB-MSCs. These findings indicate that co-administration with MIS416 enhances the therapeutic potential of hUCB-MSCs by systemically regulating the immune response, which might be an effective strategy for overcoming the current obstacles to stem cell therapy in clinical practice.

Single-Factor SOX2 Mediates Direct Neural Reprogramming of Human Mesenchymal Stem Cells via Transfection of In Vitro Transcribed mRNA.

Neural stem cells (NSCs) are a prominent cell source for understanding neural pathogenesis and for developing therapeutic applications to treat neurodegenerative disease because of their regenerative capacity and multipotency. Recently, a variety of cellular reprogramming technologies have been developed to facilitate in vitro generation of NSCs, called induced NSCs (iNSCs). However, the genetic safety aspects of established virus-based reprogramming methods have been considered, and non-integrating reprogramming methods have been developed. Reprogramming with in vitro transcribed (IVT) mRNA is one of the genetically safe reprogramming methods because exogenous mRNA temporally exists in the cell and is not integrated into the chromosome. Here, we successfully generated expandable iNSCs from human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) via transfection with IVT mRNA encoding SOX2 (SOX2 mRNA) with properly optimized conditions. We confirmed that generated human UCB-MSC-derived iNSCs (UM-iNSCs) possess characteristics of NSCs, including multipotency and self-renewal capacity. Additionally, we transfected human dermal fibroblasts (HDFs) with SOX2 mRNA. Compared with human embryonic stem cell-derived NSCs, HDFs transfected with SOX2 mRNA exhibited neural reprogramming with similar morphologies and NSC-enriched mRNA levels, but they showed limited proliferation ability. Our results demonstrated that human UCB-MSCs can be used for direct reprogramming into NSCs through transfection with IVT mRNA encoding a single factor, which provides an integration-free reprogramming tool for future therapeutic application.

GATA4-dependent regulation of the secretory phenotype via MCP-1 underlies lamin A-mediated human mesenchymal stem cell aging.

Defects in the nuclear lamina occur during physiological aging and as. result of premature aging disorders. Aging is also accompanied by an increase in transcription of genes encoding cytokines and chemokines,. phenomenon known as the senescence-associated secretory phenotype (SASP). Progerin and prelamin. trigger premature senescence and loss of function of human mesenchymal stem cells (hMSCs), but little is known about how defects in nuclear lamin. regulate SASP. Here, we show that both progerin overexpression and ZMPSTE24 depletion induce paracrine senescence, especially through the expression of monocyte chemoattractant protein-1 (MCP-1), in hMSCs. Importantly, we identified that GATA4 is. mediator regulating MCP-1 expression in response to prelamin. or progerin in hMSCs. Co-immunoprecipitation revealed that GATA4 expression is maintained due to impaired p62-mediated degradation in progerin-expressing hMSCs. Furthermore, depletion of GATA4 abrogated SASP-dependent senescence through suppression of NF-ĸB and MCP-1 in hMSCs with progerin or prelamin A. Thus, our findings indicate that abnormal lamin. proteins trigger paracrine senescence through. GATA4-dependent pathway in hMSCs. This molecular link between defective lamin. and GATA4 can provide insights into physiological aging and pathological aging disorders.

Donor-dependent variation of human umbilical cord blood mesenchymal stem cells in response to hypoxic preconditioning and amelioration of limb ischemia.

With the rapidly growing demand for mesenchymal stem cell (MSC) therapy, numerous strategies using MSCs for different diseases have been studied and reported. Because of their immunosuppressive properties, MSCs are commonly used as an allogeneic treatment. However, for the many donors who could potentially be used, it is important to understand the capacity for therapeutic usage with donor-to-donor heterogeneity. In this study, we aimed to investigate MSCs as a promising therapeutic strategy for critical limb ischemia. We evaluated MSCs from two donors (#55 and #64) and analyzed the capacity for angiogenesis through in vivo and in vitro assays to compare the therapeutic effect between different donors. We emphasized the importance of intra-population heterogeneity of MSCs on therapeutic usage by evaluating the effects of hypoxia on activating cellular angiogenesis in MSCs. The precondition of hypoxia in MSCs is known to enhance therapeutic efficacy. Our study suggests that sensitivity to hypoxic conditions is different between cells originating from different donors, and this difference affects the contribution to angiogenesis. The bioinformatics analysis of different donors under hypoxic culture conditions identified intrinsic variability in gene expression patterns and suggests alternative potential genetic factors ANGPTL4, ADM, SLC2A3, and CDON as guaranteed general indicators for further stem cell therapy.