Genetic Inactivation of CD33 in Hematopoietic Stem Cells to Enable CAR T Cell Immunotherapy for Acute Myeloid Leukemia.

The absence of cancer-restricted surface markers is a major impediment to antigen-specific immunotherapy using chimeric antigen receptor (CAR) T cells. For example, targeting the canonical myeloid marker CD33 in acute myeloid leukemia (AML) results in toxicity from destruction of normal myeloid cells. We hypothesized that a leukemia-specific antigen could be created by deleting CD33 from normal hematopoietic stem and progenitor cells (HSPCs), thereby generating a hematopoietic system resistant to CD33-targeted therapy and enabling specific targeting of AML with CAR T cells. We generated CD33-deficient human HSPCs and demonstrated normal engraftment and differentiation in immunodeficient mice. Autologous CD33 KO HSPC transplantation in rhesus macaques demonstrated long-term multilineage engraftment of gene-edited cells with normal myeloid function. CD33-deficient cells were impervious to CD33-targeting CAR T cells, allowing for efficient elimination of leukemia without myelotoxicity. These studies illuminate a novel approach to antigen-specific immunotherapy by genetically engineering the host to avoid on-target, off-tumor toxicity.

A RUNX1-FPDMM rhesus macaque model reproduces the human phenotype and predicts challenges to curative gene therapies.

Germ line loss-of-function heterozygous mutations in the RUNX1 gene cause familial platelet disorder with associated myeloid malignancies (FPDMM) characterized by thrombocytopenia and a life-long risk of hematological malignancies. Although gene therapies are being considered as promising therapeutic options, current preclinical models do not recapitulate the human phenotype and are unable to elucidate the relative fitness of mutation-corrected and RUNX1-heterozygous mutant hematopoietic stem and progenitor cells (HSPCs) in vivo long term. We generated a rhesus macaque with an FPDMM competitive repopulation model using CRISPR/Cas9 nonhomologous end joining editing in the RUNX1 gene and the AAVS1 safe-harbor control locus. We transplanted mixed populations of edited autologous HSPCs and tracked mutated allele frequencies in blood cells. In both animals, RUNX1-edited cells expanded over time compared with AAVS1-edited cells. Platelet counts remained below the normal range in the long term. Bone marrows developed megakaryocytic dysplasia similar to human FPDMM, and CD34+ HSPCs showed impaired in vitro megakaryocytic differentiation, with a striking defect in polyploidization. In conclusion, the lack of a competitive advantage for wildtype or control-edited HSPCs over RUNX1 heterozygous-mutated HSPCs long term in our preclinical model suggests that gene correction approaches for FPDMM will be challenging, particularly to reverse myelodysplastic syndrome/ acute myeloid leukemia predisposition and thrombopoietic defects.

A macaque clonal hematopoiesis model demonstrates expansion of TET2-disrupted clones and utility for testing interventions.

Individuals with age-related clonal hematopoiesis (CH) are at greater risk for hematologic malignancies and cardiovascular diseases. However, predictive preclinical animal models to recapitulate the spectrum of human CH are lacking. Through error-corrected sequencing of 56 human CH/myeloid malignancy genes, we identified natural CH driver mutations in aged rhesus macaques matching genes somatically mutated in human CH, with DNMT3A mutations being the most frequent. A CH model in young adult macaques was generated via autologous transplantation of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated gene-edited hematopoietic stem and progenitor cells (HSPCs), targeting the top human CH genes with loss-of-function (LOF) mutations. Long-term follow-up revealed reproducible and significant expansion of multiple HSPC clones with heterozygous TET2 LOF mutations, compared with minimal expansion of clones bearing other mutations. Although the blood counts of these CH macaques were normal, their bone marrows were hypercellular and myeloid-predominant. TET2-disrupted myeloid colony-forming units isolated from these animals showed a distinct hyperinflammatory gene expression profile compared with wild type. In addition, mature macrophages purified from the CH macaques showed elevated NLRP3 inflammasome activity and increased interleukin-1ß (IL-1ß) and IL-6 production. The model was used to test the impact of IL-6 blockage by tocilizumab, documenting a slowing of TET2-mutated expansion, suggesting that interruption of the IL-6 axis may remove the selective advantage of mutant HSPCs. These findings provide a model for examining the pathophysiology of CH and give insights into potential therapeutic interventions.

Prediction and validation of hematopoietic stem and progenitor cell off-target editing in transplanted rhesus macaques.

The programmable nuclease technology CRISPR-Cas9 has revolutionized gene editing in the last decade. Due to the risk of off-target editing, accurate and sensitive methods for off-target characterization are crucial prior to applying CRISPR-Cas9 therapeutically. Here, we utilized a rhesus macaque model to compare the predictive values of CIRCLE-seq, an in vitro off-target prediction method, with in silico prediction (ISP) based solely on genomic sequence comparisons. We use AmpliSeq HD error-corrected sequencing to validate off-target sites predicted by CIRCLE-seq and ISP for a CD33 guide RNA (gRNA) with thousands of off-target sites predicted by ISP and CIRCLE-seq. We found poor correlation between the sites predicted by the two methods. When almost 500 sites predicted by each method were analyzed by error-corrected sequencing of hematopoietic cells following transplantation, 19 off-target sites revealed insertion or deletion mutations. Of these sites, 8 were predicted by both methods, 8 by CIRCLE-seq only, and 3 by ISP only. The levels of cells with these off-target edits exhibited no expansion or abnormal behavior in vivo in animals followed for up to 2 years. In addition, we utilized an unbiased method termed CAST-seq to search for translocations between the on-target site and off-target sites present in animals following transplantation, detecting one specific translocation that persisted in blood cells for at least 1 year following transplantation. In conclusion, neither CIRCLE-seq or ISP predicted all sites, and a combination of careful gRNA design, followed by screening for predicted off-target sites in target cells by multiple methods, may be required for optimizing safety of clinical development.

The impact of aging on primate hematopoiesis as interrogated by clonal tracking.

Age-associated changes in hematopoietic stem and progenitor cells (HSPCs) have been carefully documented in mouse models but poorly characterized in primates and humans. To investigate clinically relevant aspects of hematopoietic aging, we compared the clonal output of thousands of genetically barcoded HSPCs in aged vs young macaques after autologous transplantation. Aged macaques showed delayed emergence of output from multipotent (MP) clones, with persistence of lineage-biased clones for many months after engraftment. In contrast to murine aging models reporting persistence of myeloid-biased HSPCs, aged macaques demonstrated persistent output from both B-cell and myeloid-biased clones. Clonal expansions of MP, myeloid-biased, and B-biased clones occurred in aged macaques, providing a potential model for human clonal hematopoiesis of indeterminate prognosis. These results suggest that long-term MP HSPC output is impaired in aged macaques, resulting in differences in the kinetics and lineage reconstitution patterns between young and aged primates in an autologous transplantation setting.

Current Strategies to Enhance Adipose Stem Cell Function: An Update.

Mesenchymal stem cells (MSCs) emerged as a promising therapeutic tool targeting a variety of inflammatory disorders due to their multiple remarkable properties, such as superior immunomodulatory function and tissue-regenerative capacity. Although bone marrow (BM) is a dominant source for adult MSCs, increasing evidence suggests that adipose tissue-derived stem cells (ASCs), which can be easily obtained at a relatively high yield, have potent therapeutic advantages comparable with BM-MSCs. Despite its outstanding benefits in pre-clinical settings, the practical efficacy of ASCs remains controversial since clinical trials with ASC application often resulted in unsatisfactory outcomes. To overcome this challenge, scientists established several strategies to generate highly functional ASCs beyond the naïve cells, including (1) pre-conditioning of ASCs with various stimulants such as inflammatory agents, (2) genetic manipulation of ASCs and (3) modification of culture conditions with three-dimensional (3D) aggregate formation and hypoxic culture. Also, exosomes and other extracellular vesicles secreted from ASCs can be applied directly to recapitulate the beneficial performance of ASCs. This review summarizes the current strategies to improve the therapeutic features of ASCs for successful clinical implementation.

Mesenchymal Stem Cell Therapy for Inflammatory Skin Diseases: Clinical Potential and Mode of Action.

Inflammatory skin disorders that cause serious deterioration of the quality of life have become one of the major public concerns. Despite their significance, there is no fundamental cure to date. Mesenchymal stem cells (MSCs) possess unique immunomodulatory properties which make them a promising tool for the treatment of various inflammatory diseases. Our recent preclinical and clinical studies have shown that MSCs can be successfully used for the treatment of atopic dermatitis (AD), one of the major inflammatory skin diseases. This observation along with similar reports from other groups revealed the efficacy and underlying mechanisms of MSCs in inflammatory dermatosis. In addition, it has been proposed that cell priming or gene transduction can be novel strategies for the development of next-generation high-efficacy MSCs for treating inflammatory skin diseases. We discuss here existing evidence that demonstrates the regulatory properties of MSCs on immune responses under inflammatory conditions.

Cathepsin S contributes to microglia-mediated olfactory dysfunction through the regulation of Cx3cl1-Cx3cr1 axis in a Niemann-Pick disease type C1 model.

Microglia can aggravate olfactory dysfunction by mediating neuronal death in the olfactory bulb (OB) of a murine model of Niemann-Pick disease type C1 (NPC1), a fatal neurodegenerative disorder accompanied by lipid trafficking defects. In this study, we focused on the crosstalk between neurons and microglia to elucidate the mechanisms underlying extensive microgliosis in the NPC1-affected brain. Microglia in the OB of NPC1 mice strongly expressed CX3C chemokine receptor 1 (Cx3cr1), a specific receptor for the neural chemokine C-X3-C motif ligand 1 (Cx3cl1). In addition, a high level of Cx3cl1 was detected in NPC1 mouse-derived CSF due to enhanced catalytic activity of Cathepsin S (Ctss), which is responsible for Cx3cl1 secretion. Notably, nasal delivery of Cx3cl1 neutralizing antibody or Ctss inhibitor could inhibit the Cx3cl1-Cx3cr1 interaction and support neuronal survival through the suppression of microglial activation, leading to an improvement in the olfactory function in NPC1 mice. Relevant in vitro experiments revealed that intracellular cholesterol accumulation could act as a strong inducer of abnormal Ctss activation and, in turn, stimulated the Cx3cl1-Cx3cr1 axis in microglia via p38 mitogen-activated protein kinase signaling. Our data address the significance of Cx3cl1-Cx3cr1 interaction in the development of microglial neurotoxicity and suggest that Ctss is a key upstream regulator. Therefore, this study contributes to a better understanding of the crosstalk between neurons and microglia in the development of the neurodegeneration and provides a new perspective for the management of olfactory deficits and other microglia-dependent neuropathies. GLIA 2016;64:2291-2305.

Human umbilical cord blood mesenchymal stem cell-derived PGE2 and TGF-ß1 alleviate atopic dermatitis by reducing mast cell degranulation.

Mesenchymal stem cell (MSC) is a promising tool for the therapy of immune disorders. However, their efficacy and mechanisms in treating allergic skin disorders are less verified. We sought to investigate the therapeutic efficacy of human umbilical cord blood-derived MSCs (hUCB-MSCs) against murine atopic dermatitis (AD) and to explore distinct mechanisms that regulate their efficacy. AD was induced in mice by the topical application of Dermatophagoides farinae. Naïve or activated-hUCB-MSCs were administered to mice, and clinical severity was determined. The subcutaneous administration of nucleotide-binding oligomerization domain 2 (NOD2)-activated hUCB-MSCs exhibited prominent protective effects against AD, and suppressed the infiltration and degranulation of mast cells (MCs). A ß-hexosaminidase assay was performed to evaluate the effect of hUCB-MSCs on MC degranulation. NOD2-activated MSCs reduced the MC degranulation via NOD2-cyclooxygenase-2 signaling. In contrast to bone marrow-derived MSCs, hUCB-MSCs exerted a cell-to-cell contact-independent suppressive effect on MC degranulation through the higher production of prostaglandin E2 (PGE2 ). Additionally, transforming growth factor (TGF)-ß1 production from hUCB-MSCs in response to interleukin-4 contributed to the attenuation of MC degranulation by downregulating FcεRI expression in MCs. In conclusion, the subcutaneous application of NOD2-activated hUCB-MSCs can efficiently ameliorate AD, and MSC-derived PGE2 and TGF-ß1 are required for the inhibition of MC degranulation.

MicroRNA-141-3p plays a role in human mesenchymal stem cell aging by directly targeting ZMPSTE24.

Human mesenchymal stem cell (hMSC) aging may lead to a reduced tissue regeneration capacity and a decline in physiological functions. However, the molecular mechanisms controlling hMSC aging in the context of prelamin A accumulation are not completely understood. In this study, we demonstrate that the accumulation of prelamin A in the nuclear envelope results in cellular senescence and potential downstream regulatory mechanisms responsible for prelamin A accumulation in hMSCs. We show for the first time that ZMPSTE24, which is involved in the post-translational maturation of lamin A, is largely responsible for the prelamin A accumulation related to cellular senescence in hMSCs. Direct binding of miR-141-3p to the 3'UTR of ZMPSTE24 transcripts was confirmed using a 3'UTR-luciferase reporter assay. We also found that miR-141-3p, which is overexpressed during senescence as a result of epigenetic regulation, is able to decrease ZMPSTE24 expression levels, and leads to an upregulation of prelamin A in hMSCs. This study provides new insights into mechanisms regulating MSC aging and may have implications for therapeutic application to reduce age-associated MSC pool exhaustion.

Bacteriophage therapy of a Vibrio parahaemolyticus infection caused by a multiple-antibiotic-resistant O3:K6 pandemic clinical strain.

BACKGROUND: Recently isolated Vibrio parahaemolyticus strains have displayed multiple antibiotic resistance. Alternatives to conventional antibiotics are needed, especially for the multiple-antibiotic-resistant V. parahaemolyticus pandemic strain. METHODS: A bacteriophage, designated pVp-1, showed effective infectivity for multiple-antibiotic-resistant V. parahaemolyticus and V. vulnificus, including V. parahaemolyticus pandemic strains. The therapeutic potential of the phage was studied in a mouse model of experimental infection using a multiple-antibiotic-resistant V. parahaemolyticus pandemic strain. We monitored the survivability and histopathological changes, quantified the bacterial and phage titers during phage therapy, and observed the immune response induced by phage induction. RESULTS: Phage-treated mice displayed protection from a V. parahaemolyticus infection and survived lethal oral and intraperitoneal bacterial challenges. CONCLUSIONS: To the best of our knowledge, this is the first report of phage therapy in a mouse model against a multiple-antibiotic­resistant V. parahaemolyticus pandemic strain infection.

Human umbilical cord blood mesenchymal stem cells reduce colitis in mice by activating NOD2 signaling to COX2.

BACKGROUND & AIMS: Decreased levels or function of nucleotide-binding oligomerization domain 2 (NOD2) are associated with Crohn's disease. NOD2 regulates intestinal inflammation, and also is expressed by human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs), to regulate their differentiation. We investigated whether NOD2 is required for the anti-inflammatory activities of MSCs in mice with colitis. METHODS: Colitis was induced in mice by administration of dextran sulfate sodium or trinitrobenzene sulfonic acid. Mice then were given intraperitoneal injections of NOD2-activated hUCB-MSCs; colon tissues and mesenteric lymph nodes were collected for histologic analyses. A bromodeoxyuridine assay was used to determine the ability of hUCB-MSCs to inhibit proliferation of human mononuclear cells in culture. RESULTS: Administration of hUCB-MSCs reduced the severity of colitis in mice. The anti-inflammatory effects of hUCB-MSCs were greatly increased by activation of NOD2 by its ligand, muramyl dipeptide (MDP). Administration of NOD2-activated hUCB-MSCs increased anti-inflammatory responses in colons of mice, such as production of interleukin (IL)-10 and infiltration by T regulatory cells, and reduced production of inflammatory cytokines. Proliferation of mononuclear cells was inhibited significantly by co-culture with hUCB-MSCs that had been stimulated with MDP. MDP induced prolonged production of prostaglandin (PG)E2 in hUCB-MSCs via the NOD2-RIP2 pathway, which suppressed proliferation of mononuclear cells derived from hUCB. PGE2 produced by hUCB-MSCs in response to MDP increased production of IL-10 and T regulatory cells. In mice, production of PGE2 by MSCs and subsequent production of IL-10 were required to reduce the severity of colitis. CONCLUSIONS: Activation of NOD2 is required for the ability of hUCB-MSCs to reduce the severity of colitis in mice. NOD2 signaling increases the ability of these cells to suppress mononuclear cell proliferation by inducing production of PGE2.

Investigation of the impact of clonal hematopoiesis on severity and pathophysiology of COVID-19 in rhesus macaques.

Clinical manifestations of COVID-19 vary widely, ranging from asymptomatic to severe respiratory failure with profound inflammation. Although risk factors for severe illness have been identified, definitive determinants remain elusive. Clonal hematopoiesis (CH), the expansion of hematopoietic stem and progenitor cells bearing acquired somatic mutations, is associated with advanced age and hyperinflammation. Given the similar age range and hyperinflammatory phenotype between frequent CH and severe COVID-19, CH could impact the risk of severe COVID-19. Human cohort studies have attempted to prove this relationship, but conclusions are conflicting. Rhesus macaques (RMs) are being utilized to test vaccines and therapeutics for COVID-19. However, RMs, even other species, have not yet been reported to develop late inflammatory COVID-19 disease. Here, RMs with either spontaneous DNMT3A or engineered TET2 CH along with similarly transplanted and conditioned controls were infected with SARS-CoV-2 and monitored until 12 days post-inoculation (dpi). Although no significant differences in clinical symptoms and blood counts were noted, an aged animal with natural DNMT3A CH died on 10 dpi. CH macaques showed evidence of sustained local inflammatory responses compared to controls. Interestingly, viral loads in respiratory tracts were higher at every timepoint in the CH group. Lung sections from euthanasia showed evidence of mild inflammation in all animals, while viral antigen was more frequently detected in the lung tissues of CH macaques even at the time of autopsy. Despite the lack of striking inflammation and serious illness, our findings suggest potential pathophysiological differences in RMs with or without CH upon SARS-CoV-2 infection.

Investigation of the Impact of Clonal Hematopoiesis on Severity and Pathophysiology of COVID-19 in Rhesus Macaques.

Clinical manifestations of COVID-19 vary widely, ranging from asymptomatic to severe respiratory failure with profound inflammation. Although risk factors for severe illness have been identified, definitive determinants remain elusive. Clonal hematopoiesis (CH), the expansion of hematopoietic stem and progenitor cells bearing acquired somatic mutations, is associated with advanced age and hyperinflammation. Given the similar age range and hyperinflammatory phenotype between frequent CH and severe COVID-19, CH could impact the risk of severe COVID-19. Human cohort studies have attempted to prove this relationship, but conclusions are conflicting. Rhesus macaques (RMs) are being utilized to test vaccines and therapeutics for COVID-19. However, RMs, even other species, have not yet been reported to develop late inflammatory COVID-19 disease. Here, RMs with either spontaneous DNMT3A or engineered TET2 CH along with similarly transplanted and conditioned controls were infected with SARS-CoV-2 and monitored until 12 days post-inoculation (dpi). Although no significant differences in clinical symptoms and blood counts were noted, an aged animal with natural DNMT3A CH died on 10 dpi. CH macaques showed evidence of sustained local inflammatory responses compared to controls. Interestingly, viral loads in respiratory tracts were higher at every timepoint in the CH group. Lung sections from euthanasia showed evidence of mild inflammation in all animals, while viral antigen was more frequently detected in the lung tissues of CH macaques even at the time of autopsy. Despite the lack of striking inflammation and serious illness, our findings suggest potential pathophysiological differences in RMs with or without CH upon SARS-CoV-2 infection. Highlights: No evidence of association between CH and COVID-19 clinical severity in macaques.The presence of CH is associated with prolonged local inflammatory responses in COVID-19.SARS-CoV-2 persists longer in respiratory tracts of macaques with CH following infection.

Mesenchymal stem cells target microglia via galectin-1 production to rescue aged mice from olfactory dysfunction.

Olfactory loss has been considered as the earliest complication for the aging process while underlying mechanisms and therapeutic strategies remain unclear. Given the correlation between microglial activation and olfactory dysfunction, here we investigated whether the immunomodulatory action of mesenchymal stem cells (MSCs) can rescue the olfactory impairment in old mice. The intranasal delivery of MSCs limited microglial activation and neuronal apoptosis in the olfactory bulb (OB), leading to improvement in olfaction. MSCs down-regulated the proportion of CD86+ microglia and prevented the maturation of cathepsin S, one of the inflammatory mediators in olfactory impairment, via the suppression of p38 MAPK signaling. Notably, old astrocytes could not prevent excessive microgliosis because the endogenous production of Galectin-1 (Gal1), one of the key microglia regulators secreted by astrocytes, was not sufficiently upregulated in the aged brain despite the presence of reactive astrogliosis. Considering that Gal1 is known as a potent paracrine factor of MSCs, we investigated whether MSC-derived Gal1 could compensate for defective astrocyte function in terms of microglial regulation. MSCs and their culture supernatant (MSC-CM) could regulate the direction of microglial differentiation by impeding the polarization towards the pro-inflammatory M1 type; notably, a selective Gal1 inhibitor OTX008 could hinder this phenomenon, indicating that Gal1 is involved in immunomodulation exerted by MSCs. Also, acute microglial activation within the OB upon LPS infusion was attenuated by MSC-CM in a Gal1-dependent manner. Our study demonstrates the therapeutic benefit of MSCs on age-related olfactory dysfunction and suggests Gal1 as a key mediator of the anti-inflammatory action of MSCs.

Respiratory and Systemic Toxicity of Inhaled Artificial Asian Sand Dust in Pigs.

Air pollution, particularly caused by Asian sand dust (ASD) and particulate matter (PM), has become one of the leading threats to public health. However, the majority of studies have primarily focused on epidemiological assessment, and in vivo toxicities of certain air pollutants have been poorly elucidated in medium/large-size laboratory animals. To investigate the impact of ASD in domestic animals, 16 Landrace pigs were exposed to an artificial ASD sandstorm for 6 h. All animals were divided in four cages, and a commercial yellow soil was used for generating artificial mineralogical particles. Blood samples were collected, and necropsies were performed before exposure and 6, 12, 24, and 72 h after exposure. Complete blood cell count and the levels of serum biochemical enzymes, blood gas, electrolytes, and a variety of inflammatory cytokines were evaluated. In addition, histopathological examination was conducted. Various test results proved acute lower airway disorders with systemic inflammation in pigs. To our knowledge, this study is the first to describe experimental research in domestic animals concerning the damage caused by artificial ASD exposure. The results of this study suggest that ASD has importance in terms of not only public health but also of ultimate economic losses in the pork industry.

TNF-α Priming Elicits Robust Immunomodulatory Potential of Human Tonsil-Derived Mesenchymal Stem Cells to Alleviate Murine Colitis.

Mesenchymal stem cells (MSCs) have been spotlighted in the field of cell therapies as a promising tool for the treatment of intractable inflammatory diseases. However, their therapeutic potency still shows a gap between preclinical and clinical settings, and distinctive characteristics of specific tissue-derived MSCs and definitive ways to maximize their beneficial functions have not been fully elucidated yet. We previously identified the unique MSCs population from human palatine tonsil (TMSCs) and revealed their superior properties in proliferation and ROS regulation. Based on these findings, we explored further characteristics of TMSCs particularly focused on immunomodulatory function. We found the merit of TMSCs as a therapeutic agent that retains favorable MSCs properties until relatively late passages and revealed that pre-treatment of TNF-α can enhance the immunomodulatory abilities of TMSCs through the upregulation of the PTGS2/PGE2 axis. TMSCs primed with TNF-α effectively restrained the proliferation and differentiation of T lymphocytes and macrophages in vitro, and more interestingly, these TNF-α-licensed TMSCs exhibited significant prophylactic and therapeutic efficacy in a murine model of autoimmune-mediated acute colitis via clinical and histopathological assessment compared to unprimed naïve TMSCs. These findings provide novel insight into the optimization and standardization of MSCs-based anti-inflammatory therapies, especially targeting inflammatory bowel disease (IBD).

Extracellular Vesicles from SOD3-Transduced Stem Cells Exhibit Improved Immunomodulatory Abilities in the Murine Dermatitis Model.

The immunoregulatory abilities of mesenchymal stem cells (MSCs) have been investigated in various autoimmune and allergic diseases. However, the therapeutic benefits observed in preclinical settings have not been reproducible in clinical trials. This discrepancy is due to insufficient efficacy of MSCs in harsh microenvironments, as well as batch-dependent variability in potency. Therefore, to achieve more beneficial and uniform outcomes, novel strategies are required to potentiate the therapeutic effect of MSCs. One of simple strategies to augment cellular function is genetic manipulation. Several studies showed that transduction of antioxidant enzyme into cells can increase anti-inflammatory effects. Therefore, we evaluated the immunoregulatory abilities of MSCs introduced with extracellular superoxide dismutase 3 (SOD3) in the present study. SOD3-overexpressed MSCs (SOD3-MSCs) reduced the symptoms of murine model of atopic dermatitis (AD)-like inflammation, as well as the differentiation and activation of various immune cells involved in AD progression. Interestingly, extracellular vesicles (EVs) isolated from SOD3-MSCs delivered SOD3 protein. EVs carrying SOD3 also exerted improved therapeutic efficacy, as observed in their parent cells. These results suggest that MSCs transduced with SOD3, an antioxidant enzyme, as well as EVs isolated from modified cells, might be developed as a promising cell-based therapeutics for inflammatory disorders.